text:
Tour de Taiwan - Stage 4 in Nantou County on March 20
2019 Tour de Taiwan - Stage 4 in Nantou County kicked off grandly at the County government office at 9:00 am on March 20. 181 international outstanding riders flew to Nantou from 5 continents for the yellow champion jersey. Ming-Chen Lin, the Magistrate of Nantou County, said the route distance is about 170km and is mainly mountain. Riders would circle around Nantou 1.5 times and would see a lot of beautiful landscapes. We expect to promote Taiwan's publicity on the international platform through international events.
Jonathan Clarke (AUS) of Floyd's Pro Cycling from Canada snagged the individual championship (yellow jersey), and the green jersey (King of Sprint) went to Yasuharu Nakajima (JPN) of Kinan Cycling Team from Japan. The Polka Dot Jersey (King of the Mountains) was given to Wilmar Andres Paredes Zapata (COL) of Manzana Postobon Team from Columbia. James Piccoli of Elevate-KHS Pro Cycling Team from USA made a hectic sprint and won the queen stage. Feng, Chun-Kai of Chinese Taipei Cycling Team kept the blue jersey (Best Asian Rider) in Taiwan.


December 2010 - Deb Arnold, Ink.
It's the holidays, after all.
So, I guess I've just been inspired by Will Smith and family lately. In wishing to tell you about my awesome holiday video, "Getting Jiggy Wit It," Will's 1997 hit, (forgive the likely-illegal link; finding 14-year-old videos is a challenge) was suddenly on my lips.
Please note that the entire video was shot on an iPhone 4, and edited on a Mac – all produced by Steven Neuman Design.
I think you'll agree that Steven is a *genius*.
Andrew McAfee was the first to name it back in March 2006: the then-new phenomenon of companies leveraging Web 2.0 platforms and technologies inside the business – as a means to connect employees, make processes more efficient and/or connect with partners and customers.
At the recent Enterprise 2.0 Conference in Santa Clara, there was, despite the event's name, hot debate about whether the phenomenon should be called "social business." Those in favor contrasted sharply with speakers cautioning us in the audience about using the word "social" at all, as it tends to spook the C-level.
The article, "The rise of the networked enterprise: Web 2.0 finds its payday" indicates that "fully networked enterprises," those successful at leveraging Web 2.0 both internally and externally, note improvements in market share gains, operating margins and market leadership.
Blogs, mash-ups (a Web application combines multiple sources of data into a single tool), microblogging, peer to peer, podcasts, prediction markets, rating, RSS (Really Simple Syndication), social networking, tagging, video sharing, and wikis.
Now, I'm not sure I would go so far as to say that if McKinsey says it, it must be true; however, I think it's fair to say that it's worth paying close attention.
Which I intend to keep doing.
Now, I don't typically write about pop culture but I was recently introduced to the band OK Go and their truly amazing videos (thanks to Steven Neuman, who helped me create a pretty sweet video of my own…to come).
Fairly bare-bones sets (with some exceptions; see below) and the judicious use of choreography and movement.
Very, very different from another video I've been watching non-stop, as my friends can attest: Whip My Hair by Willow Smith, daughter of Will and Jada Pinkett Smith. She's my new female role model. She just turned 10.
Whip My Hair, i.e. be my true self.
Her video is very produced, very complex, very high-tech (it helps to have friends in high places). Basically a three-minute wave of color, movement and sound.
And DANG that girl's got attitude. The right kind. The very best kind.
So for contrast, check out these OK Go videos. I hereby solemnly swear that they will be worth your time.
– Here It Goes Again – homemade, starring the four guys in the band and six treadmills. Clever.
– White Knuckles – starring the four guys in the band and a slew of trained dogs. Hilarious.
– This Too Shall Pass – amazingly complex Rube Goldberg machine that apparently required 48 takes before each intricate step worked. Genius.
What I appreciate about art is that it makes me reconsider, in some way, how to view the world and its possibilities. These videos accomplish that.


The 14 meter long fishing vessel Bøfjord caught fire near Averøy, Norway. The Bøfjord was headed to Lofoten from Ålesund when the vessel caught fire. The two crew alerted authorities who dispatched a rescue boat to the scene. The rescue boat rescued the two crew of the Bøfjord and took the fishing vessel under tow to Averøy. There the local fire brigade boarded the Bøfjord and brought the fire under control. No reports of injuries.
On January 28, the fishing boat Miss Lynn burned and sank off Port Clyde, Maine. The crew on the Miss Lynn alerted authorities that a fire had broken out in the vessel's engine room. The Coast Guard dispatched a response boat along with the Cutter Willow to the scene. Before authorities arrived, the two crew on board donned survival suits and jumped into the sea to escape the blaze. A nearby fishing vessel Bug Catcha rescued the two crew. No reports of injuries. The Cutter Willow arrived on scene, but was unable to extinguish the blaze on board the Miss Lynn. The Miss Lynn sank a short time later off Cilley Ledge.
On January 23, the 50 foot long fishing vessel Tamara became disabled off Block Island, Rhode Island. The Tamara requested assistance after its navigation equipment had failed while in strong winter storm with winds reaching 55 knots and 4 meter high waves. The Coast Guard dispatched a response boat, motor lifeboat and helicopter to the scene. The response boat arrived on scene and was able to attach a towline to the Tamara. Both vessels proceeded to Newport and arrived later the same day. No reports of injuries, damage or pollution released.


text:
Research ArticleFragile X Syndrome
Sustained correction of associative learning deficits after brief, early treatment in a rat model of Fragile X Syndrome
View ORCID ProfileAntonis Asiminas1,2,3,*,
View ORCID ProfileAdam D. Jackson1,2,3,4,*,
Susana R. Louros1,2,3,†,
Sally M. Till1,2,3,†,
View ORCID ProfileTeresa Spano1,2,3,4,
View ORCID ProfileOwen Dando1,2,3,5,
View ORCID ProfileMark F. Bear6,
Sumantra Chattarji2,3,4,
View ORCID ProfileGiles E. Hardingham1,2,3,5,
View ORCID ProfileEmily K. Osterweil1,2,3,‡,
View ORCID ProfileDavid J. A. Wyllie1,2,3,4,‡,
View ORCID ProfileEmma R. Wood1,2,3,4,§,|| and
View ORCID ProfilePeter C. Kind1,2,3,4,§,||
1Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
2Simons Initiative for the Developing Brain, University of Edinburgh, Edinburgh EH8 9XD, UK.
3Patrick Wild Centre, University of Edinburgh, Edinburgh EH8 9XD, UK.
4Centre for Brain Development and Repair, InStem, Bangalore 560065, India.
5UK Dementia Research Institute at the Edinburgh Medical School, University of Edinburgh, Edinburgh EH8 9XD, UK.
6Department of Brain and Cognitive Sciences, Howard Hughes Medical Institute, Picower Institute for Learning and Memory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
↵||Corresponding author. Email: p.kind{at}ed.ac.uk (P.C.K.); emma.wood{at}ed.ac.uk (E.R.W.)
↵† These authors contributed equally to this work.
↵‡ These authors contributed equally to this work.
↵§ These authors contributed equally to this work.
Science Translational Medicine 29 May 2019:
Vol. 11, Issue 494, eaao0498
DOI: 10.1126/scitranslmed.aao0498
Antonis Asiminas
ORCID record for Antonis Asiminas
Adam D. Jackson
ORCID record for Adam D. Jackson
Susana R. Louros
Sally M. Till
Teresa Spano
ORCID record for Teresa Spano
Owen Dando
ORCID record for Owen Dando
Mark F. Bear
ORCID record for Mark F. Bear
Sumantra Chattarji
Giles E. Hardingham
ORCID record for Giles E. Hardingham
Emily K. Osterweil
ORCID record for Emily K. Osterweil
David J. A. Wyllie
ORCID record for David J. A. Wyllie
Emma R. Wood
ORCID record for Emma R. Wood
For correspondence: p.kind@ed.ac.uk emma.wood@ed.ac.uk
Peter C. Kind
ORCID record for Peter C. Kind
Optimizing therapeutic intervention for Fragile X Syndrome
Fragile X syndrome (FXS) is a neurodevelopmental disorder caused by mutations in the FMR1 gene. Pharmacological intervention using the cholesterol-lowering drug lovastatin had therapeutic effect on behavior in patients. Selecting the time of intervention and the therapeutic protocol is critical for maximizing the therapeutic effects. Now, Asiminas et al. tested the effect of temporary early treatment with lovastatin in a rat model of FXS. The authors showed that 5-week treatment initiated before complete development of cognitive abilities rescued cognitive development. The therapeutic effect persisted for more than 3 months after treatment termination. The results suggest that lovastatin treatment initiated early during development might have disease-modifying effect in FXS.
Fragile X Syndrome (FXS) is one of the most common monogenic forms of autism and intellectual disability. Preclinical studies in animal models have highlighted the potential of pharmaceutical intervention strategies for alleviating the symptoms of FXS. However, whether treatment strategies can be tailored to developmental time windows that define the emergence of particular phenotypes is unknown. Similarly, whether a brief, early intervention can have long-lasting beneficial effects, even after treatment cessation, is also unknown. To address these questions, we first examined the developmental profile for the acquisition of associative learning in a rat model of FXS. Associative memory was tested using a range of behavioral paradigms that rely on an animal's innate tendency to explore novelty. Fmr1 knockout (KO) rats showed a developmental delay in their acquisition of object-place recognition and did not demonstrate object-place-context recognition paradigm at any age tested (up to 23 weeks of age). Treatment of Fmr1 KO rats with lovastatin between 5 and 9 weeks of age, during the normal developmental period that this associative memory capability is established, prevents the emergence of deficits but has no effect in wild-type animals. Moreover, we observe no regression of cognitive performance in the FXS rats over several months after treatment. This restoration of the normal developmental trajectory of cognitive function is associated with the sustained rescue of both synaptic plasticity and altered protein synthesis. The findings provide proof of concept that the impaired emergence of the cognitive repertoire in neurodevelopmental disorders may be prevented by brief, early pharmacological intervention.
Fragile X Syndrome (FXS) is a major heritable cause of intellectual disability and one of the most common single-gene causes of autism spectrum disorder (ASD), with 30 to 50% of boys clinically diagnosed with ASD (1). It affects about 1:4000 boys and 1:6000 to 8000 girls. FXS has numerous co-occurring conditions including anxiety disorders, sensory hypersensitivity, and seizures (1). FXS is usually diagnosed around 3 years of age as a result of a delay in language development (2). However, early diagnosis through genetic screening suggests early symptom development in agreement with data from cellular phenotypes in rodent models (2–4). FXS is typically caused by an expansion of a trinucleotide repeat (CGG) in the promoter region of the FMR1 gene that leads to silencing of the gene and no protein expression (5), although de novo mutations that are predicted to alter protein function also cause FXS (6, 7).
There is abundant preclinical evidence that an array of functional impairments in FXS arise from a disruption of cellular biochemistry and physiology that is correctable with pharmacological interventions [for reviews see (2, 8–10)]. Furthermore, on the basis of knowledge of critical periods in sensory system development and language acquisition (11, 12), it has been suggested that therapeutic success would be improved by starting treatments early, before major symptoms develop (13). However, it is unknown whether effective treatments would need to be maintained throughout life.
Numerous clinical trials have taken place for FXS using newly developed compound or repurposed drugs (14). There are numerous advantages of drug repurposing in the treatment of disease, including speed of translation and cost implications associated with new drug development (15). When assessing the feasibility of initiating chronic treatments in infancy, an obvious concern is safety. For these reasons, there has been interest in the possibility of repurposing drugs with a known safety profile in children. Two examples of such candidate drugs are arbaclofen, a γ-aminobutyric acid type B (GABAB) receptor agonist used for the treatment of spasticity in cerebral palsy, and lovastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor used for the treatment of hypercholesterolemia. These compounds have been shown to correct the alterations in synaptic plasticity, neuronal morphology, neuronal excitability and behavior in a mouse model of FXS (16, 17). The precise mechanisms underlying this rescue in Fmr1 knockout (KO) mice are not known; however, increasing GABAB activation with arbaclofen alters the excitability of neurons and, via the inhibition of glutamate release, reduces the activation of the metabotropic glutamate receptor (mGluR) 5 (18)—two key cellular processes thought to underlie FXS pathophysiology (9). Lovastatin regulates the membrane association of Ras and subsequently mildly reduces the downstream activation of the extracellular signal–regulated kinases (ERKs) 1 and 2 (17). A recent human study evaluating the effects of arbaclofen in children and adolescents with FXS suggested that age may indeed be a critical variable in determining the effect of the treatment; arbaclofen had therapeutic effects in children that were lost in adolescent/adults with FXS (19). Open-label phase 2 clinical trials with lovastatin in FXS were promising (20); follow-up studies have not been reported yet (NCT02680379, NCT02998151, and NCT02642653). The combined age range in these trials is 8 to 55 years of age.
As noted above, a key question for the potential treatment of FXS is the age at which treatment should begin to maximize effectiveness. In this study, we tested the hypothesis that lovastatin treatment restricted to early development could permanently prevent impairments in cognitive function in an animal model of FXS. We chose lovastatin because it can be orally administered in food, has been shown to ameliorate circuit deficits in Fmr1 KO mice, and has a known safety profile in children (17).
Key to addressing this question is knowledge of the development of cognitive abilities in animal models. Robust behavioral paradigms independent from reward-based learning that can be used repeatedly in the same animals are therefore required. Therefore, to assess cognition across juvenile development in rats, we used a battery of spontaneous object exploration "novelty preference" tasks. These tasks are based on rodents' innate preference to explore novel over familiar stimuli in their environment (21) and can be used to infer memory for the familiar stimulus (22). The four task variants used in the current study test the ability to discriminate between novel and familiar objects [object recognition (OR)] (22); novel and familiar configurations of objects and local contextual cues within the testing box [object-context recognition (OCR)] (23, 24); novel and familiar configurations of objects and their spatial locations within the testing box [object-place recognition (OPR)] (25); or novel and familiar configurations of objects, their spatial locations, and the local context [object-place-context recognition (OPCR)] (25). These tasks were chosen for several reasons. First, we have previously reported that, whereas adult Fmr1 KO rats show intact short-term recognition (2 to 5 min) in the OR, OCR, and OPR tasks, unlike their wild-type (WT) littermates, they do not show intact short-term memory in the OPCR task (26). Therefore, we could assess whether treatment during juvenile development would ameliorate the deficits in OPC memory in Fmr1 KO rats. Second, the tasks are based on spontaneous behavior and do not depend on the acquisition of task rules and stimulus-reward contingencies. This means that the same animal can be tested on multiple occasions (with different objects) to assess recognition memory at different ages. Third, the spontaneous exploration protocol has been used successfully in juvenile rats to characterize the ontogeny of recognition memory abilities in rats. These studies have shown that OR memory emerges before postnatal day 17 (P17) in the Long Evans Hooded (LEH) rat (27), whereas OCR emerges at around P17 or P26, depending on the types of cues used to define context (28). In contrast, OPR (29) and OPCR (30) emerge later during juvenile development. Last, short-term (2 to 5 min) recognition memory in the four different task variants described above is thought to depend on different overlapping brain circuits (31). Whereas the complete circuitry supporting recognition memory in each task is not fully resolved, it is generally agreed that the perirhinal cortex is required for OR, whereas the entorhinal cortex, hippocampus, and medial prefrontal cortex (mPFC) are not (although there is debate concerning the role of the hippocampus at longer retention intervals and its normal role in the intact brain) (23–25, 31–34). For the version of the OCR task described here (in which context is signaled by local nonpolarizing cues), the postrhinal cortex and lateral entorhinal cortex (LEC) are necessary (24, 34, 35). The LEC, together with its connections with the mPFC, is required for memory in the OPR task (33, 34), whereas for the OPCR task, the hippocampus is required, together with the LEC and its connections with mPFC (25, 32, 33).
In this study, we examine the developmental emergence of associative memory using these object exploration tasks in a rat model of FXS. We then examine whether deficits in associative memory can be prevented by early treatment with lovastatin and whether any benefits of treatment persist after treatment termination. Last, we examine whether lovastatin treatment corrects alterations at cellular levels in hippocampus and mPFC, two regions known to be involved in the tasks used in the study and known to have altered function in FXS (36, 37).
WT LEH rats show distinct developmental trajectories in different spontaneous object exploration tasks
As the first step to determining the emergence of cognitive abilities in rats, we conducted a longitudinal study to examine the developmental time course over which WT LEH rats exhibit nonassociative and associative recognition memory capabilities by examining their performance on a battery of four spontaneous exploration-based tasks from 4 weeks old to adulthood. In all tasks, memory is inferred on the basis of the amount of time that an animal spends, preferentially exploring the novel object or novelty based on an object's location and/or context (Fig. 1A). At 4 to 6 weeks old, rats exhibit novelty preference in both the nonassociative OR task and the associative OCR task (Fig. 1, B and C), indicating memory of objects and object-context associations. Memory was observed at all subsequent ages tested for these tasks. In contrast, novelty preference was not observed in the associative OPR and OPCR tasks at 4 to 6 weeks of age but was present at 7 to 9 weeks and at later ages (Fig. 1, D and E). This indicates that the circuits necessary for these associative memory processes are not fully mature until 7 weeks of age. Total exploration time did not differ between time points for any task (Fig. 1, B to E).
Fig. 1 WT LEH rats show distinct developmental trajectories in different spontaneous object exploration tasks.
(A) Schematic of the spontaneous object exploration tasks used. S, sample phase; T, test phase. Light and dark gray backgrounds denote distinct contexts, and orange symbols denote novel object/object association. Discrimination index (DI) over time for WT LEH rats in (B) OR, (C) OCR, (D) OPR, and (E) OPCR tasks. Insets (B to E) Total object exploration time during the test phase for each task. For all tasks, n(4–6) = 13, n(7–9) = 13, n(>10) = 11; *P < 0.05 difference from chance (DI = 0) based on one-sample t tests. P values from one-sample t tests have been corrected for false discovery rate using the Benjamini-Hochberg procedure. For details on t, df, and P values for one-sample t tests, see table S1.
Loss of FMRP leads to selective deficits in OPR and OPCR memory in Fmr1 KO rats
We generated an Fmr1 KO rat line on the outbred LEH strain (Fig. 2A) to determine whether the developmental trajectory of associative memory was altered as a result of chronic deletion of the Fmr1 gene. In Fmr1 KO LEH rats, no Fmr1 transcripts were detected by reverse transcription polymerase chain reaction (RT-PCR) (Fig. 2B). This was also confirmed by RNA sequencing. Similarly, no protein was detected using either Western blotting or immunohistochemistry (Fig. 2, C and D), confirming the complete absence of Fragile X protein (FMRP). In agreement with previous findings in the mouse (33) and Sprague-Dawley (SD) rats (26), FMRP is expressed throughout postnatal development in WT LEH rats (Fig. 2E).
Fig. 2 Loss of FMRP leads to selective deficits in OPR and OPCR memory in Fmr1 KO rats.
(A) Zinc finger nuclease (ZFN)–mediated disruption of Fmr1. Diagrams illustrate the target site for ZFN cleavage, donor DNA sequence including enhanced green fluorescent protein (eGFP) and a nuclear localization signal flanked by 5′ and 3′ homology recombination arms for homology-directed repair, and the resulting targeted locus (top). 5′UTR, 5′ untranslated region. (B) RT-PCR for Fmr1 and eGFP mRNA in WT and Fmr1 KO rats. Lanes 1 to 3, samples from three WT rats; lanes 4 to 6, samples from three Fmr1 KO rats; lane 7, RT control from one Fmr1 KO rat; lane 8, GFP-positive control. cDNA, complementary DNA. (C) Western blot analyses of FMRP and GFP expression in WT and Fmr1 KO rats. Lanes 1 to 2, samples from two WT rats; lanes 3 to 5, samples from three Fmr1 KO rats; lane 6, positive control for GFP. (D) Immunohistochemical localization of FMRP in P15 WT and Fmr1 KO rats. Scale bar, 500 μm. (E) Western blot analysis of FMRP expression in hippocampal homogenates from WT littermates over postnatal development compared with P15 Fmr1 KO rat. DI at different ages for WT and Fmr1 KO rats in (F) OR, (G) OCR, (H) OPR, and (I) OPCR tasks. For all tasks, nWT(4–6) = 13, nWT(7–9) = 13, nWT(>10) = 11, nKO(4–6) = 12, nKO(7–9) = 12, nKO(>10) = 11; *P< 0.05 difference from chance (DI = 0), black for WT and red for KO; #P< 0.05 difference between groups. Linear mixed effect (LME) models were fitted to the data (for details, see tables S7 and S8). P values from one-sample t tests and post hoc two-sample t tests have been controlled for the false discovery rate using the Benjamini-Hochberg procedure. For details on t, df, and P values for one-sample t tests and post hoc two-sample t tests, see table S1.
At a behavioral level, our results showed that adult (3 to 4 months old) Fmr1 KO rats on the LEH background had selective deficits in the OPCR task but intact novelty preference in the OR, OCR, and OPR tasks (fig. S1). This agrees with our previous findings in Fmr1 KO SD rats (26), indicating that this pattern of cognitive deficits is shared across outbred strains of adult rats.
Repeated testing during juvenile development (4 to 23 weeks) revealed an emergence of OR and OCR ability in Fmr1 KO LEH rats indistinguishable from that seen in WT animals (Fig. 2, F and G). In contrast, the ability of Fmr1 KO rats to show significant memory in OPR was delayed by 2 to 4 weeks compared to WT rats (Fig. 2H). As predicted from our findings in adult rats, as shown in fig. S1, (26), the Fmr1 KO rats did not exhibit OPCR memory at any age (Fig. 2I). For each of the four tasks, the total object exploration time in the sample and test phases did not differ between genotypes across different time points (fig. S2), indicating that the deficits were not secondary to changes in motivation or ability to explore objects.
Transient treatment with lovastatin restores WT-like developmental trajectory of OPR and OPCR memory in Fmr1 KO rats and has sustained effects on memory
Previous studies in the mouse model of FXS have shown that continuous, chronic treatments can correct behavioral phenotypes such as hyperactivity in an open field (17, 38), susceptibility to audiogenic seizures (16, 17), and impairments in startle response and inhibitory avoidance (38). However, the effects of a transient treatment, initiated early in postnatal life, on cognitive impairments are still mostly unexplored. We next tested whether a transient administration of the drug lovastatin to juvenile rats would correct the cognitive deficits described above. Lovastatin was chosen for three reasons. First, by inhibiting the ERK signaling pathway (fig. S3), lovastatin has been shown to correct the excessive protein synthesis, which is a core pathophysiology of FXS, and numerous cellular and circuit-based phenotypes associated with the loss of FMRP in mice. Second, lovastatin is a U.S. Food and Drug Administration–approved drug for the treatment of a familial form of hypercholesterolaemia (39) and has a known safety profile in children. Third, lovastatin can easily be administered through the diet (17).
We tested the effect of administering a 5-week oral free-feeding laboratory chow protocol, with and without lovastatin (100 mg/kg), initiated at 4 weeks, before the emergence of associative recognition in the OPR and OPCR tasks in WT rats (Fig. 3A). Treatment condition (lovastatin versus control diet) had no effect on food intake, and weight gain did not differ between genotypes or treatment conditions (fig. S4). Moreover, the total object exploration time did not differ between experimental groups (fig. S5). However, the lovastatin treatment restored the normal developmental emergence of OPR and OPCR abilities in Fmr1 KO rats without altering that of WT rats (Fig. 3, B and C). At 7 to 9 weeks of age, both WT and Fmr1 KO animals on lovastatin showed novelty preference similar to WT animals on control chow in the OPR and OPCR tasks, whereas Fmr1 KO animals on control chow showed no evidence of memory on these tasks at that age (Fig. 3D). OR and OCR abilities, which are not impaired in the Fmr1 KO rats, were not affected by the lovastatin treatment in either genotype (fig. S6). Next, to determine whether the ability of Fmr1 KO rats to perform OPCR requires continuous exposure to lovastatin, we terminated the treatment at 9 weeks of age and tested the rats again at 14 and 23 weeks of age. Treated Fmr1 KO rats maintained their ability to perform the OPCR task even at 14 and 23 weeks of age, 5 and 14 weeks after termination of lovastatin treatment, respectively (Fig. 3E). Moreover, the animals' ability to perform the OPR remained unaffected after removal of lovastatin (see Fig. 3B). These results suggest that the restoration of the developmental trajectory of associative recognition performance by lovastatin treatment persists long after treatment termination.
Fig. 3 Transient treatment with lovastatin restores WT-like developmental trajectory of OPR and OPCR memory in Fmr1 KO rats and has sustained effects on both memory and cellular pathophysiology.
(A) Experimental time course for longitudinal assessment of cognitive development in WT and Fmr1 KO rats treated with lovastatin between 4 and 9 weeks of age. DI at different ages for WT and Fmr1 KO rats treated with control or lovastatin diet in (B) OPR and (C) OPCR tasks. (D) Effect of lovastatin treatment on OPR (left) and OPCR (right) memory tested at 7 to 9 weeks of age in WT and Fmr1 KO rats. (E) Effect of lovastatin treatment on OPCR memory in WT and Fmr1 KO rats tested at 14 and 23 weeks of age. (F) Effect of lovastatin treatment on hippocampal basal protein synthesis levels in WT and Fmr1 KO rats measured at 24 weeks of age, after behavioral testing was complete. Sample sizes: OPR 4 to 6 weeks: nWTcontrol = 13, nWTlova = 12, nKOcontrol = 12, nKOlova = 12; OPR 7 to 9 weeks: nWTcontrol = 13, nWTlova = 12, nKOcontrol = 12, nKOlova = 12; OPR 14 weeks: nWTcontrol = 10, nWTlova = 11, nKOcontrol = 11, nKOlova = 8; OPR 23 weeks: nWTcontrol = 11, nWTlova = 10, nKOcontrol = 11, nKOlova = 7; OPCR 4 to 6 weeks: nWTcontrol = 13, nWTlova = 12, nKOcontrol = 12, nKOlova = 12; OPCR 7 to 9 weeks: nWTcontrol = 13, nWTlova = 12, nKOcontrol = 12, nKOlova = 12; OPCR 14 weeks: nWTcontrol = 11, nWTlova = 11, nKOcontrol = 11, nKOlova = 8; OPCR 23 weeks: nWTcontrol = 10, nWTlova = 10, nKOcontrol = 11, nKOlova = 8; for protein synthesis, n = 6 for all groups. *P< 0.05 difference from chance (DI = 0), black for WT and red for KO; #P< 0.05 difference between groups. LMEs were fitted to the behavioral data (for details, see tables S7 and S9), and two-way analysis of variance (ANOVA) with post hoc two-sample t tests was used to analyze the effect of lovastatin on hippocampal protein synthesis levels (for details, see table S6F). P values from one-sample t tests and post hoc two-sample t tests have been controlled for the false discovery rate using the Benjamini-Hochberg procedure. For details on behavioral data t, df, and P values for one-sample t tests, see table S3, and for post hoc two-sample t tests, see table S4.
Early, brief lovastatin treatment corrects increased basal protein synthesis levels in Fmr1 KO hippocampus, 15 weeks after drug removal
We next tested whether the same brief lovastatin exposure produced sustained correction of the elevated protein synthesis that is known to result from the loss of FMRP (40–42). Because OPCR is a hippocampus-dependent task (32), we examined basal protein synthesis in hippocampal slices from the rats used in our behavioral study, harvested at 24 weeks of age. Fmr1 KO rats on control diet showed an increase in basal protein synthesis relative to WT rats even at this old age. The temporary lovastatin treatment between 4 and 9 weeks of age corrected basal protein synthesis in Fmr1 KO rats without affecting WT rats (Fig. 3F).
Lovastatin prevents the emergence of prefrontal cortex plasticity deficits associated with the loss of FMRP
We next tested whether the same regimen of brief, early lovastatin treatment could prevent the emergence of an age-dependent phenotype in synaptic plasticity associated with the loss of FMRP (43). As mPFC connections are required for OPCR [referred to by Chao et al. (33) as OCP task], and Fmr1 KO mice exhibit an age-dependent deficit in long-term potentiation (LTP) of synaptic responses in this region (43), we assessed LTP in the mPFC of Fmr1 KO rats. Our results show that, whereas LTP at layers 2 to 5 synapses of mPFC was not affected by the loss of FMRP at 4 to 6 weeks of age (Fig. 4A), LTP in Fmr1 KO rats was impaired at 10 to 12 weeks of age (Fig. 4B). Lovastatin treatment from 5 weeks of age prevented the emergence of this LTP deficit as measured in 10- to 12-week-old animals (Fig. 4C).
Fig. 4 Lovastatin prevents the emergence of plasticity deficits associated with the loss of FMRP.
(A) Left: Time course plotting field excitatory postsynaptic potential (fEPSP) slopes normalized to baseline after LTP induction in layers 2 to 5 synapses in the prelimbic mPFC taken from 4- to 6-week-old WT and Fmr1 KO rats. Right: Averages of fEPSP slopes normalized to baseline during the last 20 min of the recording (70 to 90 min). (B) Left: Time course plotting fEPSP slopes normalized to baseline after LTP induction at synapses from layer 2/3 inputs onto layer 5 neurons in the prelimbic mPFC taken from 10- to 12-week-old WT and Fmr1 KO rats. Right: Averages of fEPSP slopes normalized to baseline during the last 20 min of the recording (70 to 90 min). (C) Left: Experimental time course for assessment of the effect of lovastatin (lova) treatment beginning at 5 weeks of age on WT and Fmr1 KO plasticity in the prelimbic mPFC. Middle: Time course plotting averages of fEPSP slopes normalized to baseline after LTP induction in layers 2 to 5 synapses in prelimbic mPFC slices taken from 10- to 12-week-old WT and Fmr1 KO treated with either control or lovastatin diet. Right: Averages of fEPSP slopes normalized to baseline during the last 20 min of recordings (70 to 90 min). Insets: Example traces showing synaptic responses during baseline (black trace) and at 80 to 90 min (gray trace). Scale bar, 0.5 mV; 5 ms. Sample sizes: LTP 4 to 6 weeks, n = 6 for WT and KO; LTP 10 to 12 weeks, n = 7 for WT and KO; for lovastatin effects on LTP, nWTcontrol = 9, nWTlova = 7, nKOcontrol = 9, nKOlova = 8; *P < 0.05; **P < 0.01 difference between groups; two-way ANOVA with post hoc two-sample t tests was used for data analyses (for details, see table S6, G and H). P values for post hoc two-sample t tests have been controlled for the false discovery rate using the Benjamini-Hochberg procedure.
Lovastatin has previously been shown to rescue the exaggerated mGluR-dependent long-term depression (LTD) found in mouse hippocampus at 1 month of age (17). To determine whether a similar effect on hippocampal LTD was present in our animals, we first examined whether LTD was exaggerated at 4 to 5 weeks and at 9 to 10 weeks (the age at which lovastatin is terminated in our experiments). Although we confirmed that 3,5-dihydroxyphenylglycine (DHPG)–induced LTD is exaggerated in Fmr1 KO LEH rats at 4 to 5 weeks of age, no difference was found at 9 to 10 weeks. Therefore, the effects of lovastatin treatment in our paradigm could not be examined. These data suggest that the downstream mechanisms supporting mGluR-dependent LTD change during postnatal development (fig. S7).
This study sought to test the hypothesis that early and transient therapeutic intervention can produce long-lasting benefits on cognition in a rat model of FXS. Our results demonstrate that early lovastatin treatment for 5 weeks, initiated before OPR and OPCR capabilities emerge, restored the normal developmental trajectory of these cognitive abilities in Fmr1 KO rats. Furthermore, the ability to perform associative recognition in these tasks persisted for at least 14 weeks after the end of treatment. These findings indicate that brief, early treatment not only prevents the emergence of cognitive deficits but also these beneficial effects on cognition are sustained long after the end of treatment. They also suggest that lovastatin treatment during an early developmental time window might rescue the normal development of the neural circuits underlying these behaviors and that FMRP, lacking in these animals, might not be needed for their maintenance.
Natural history studies of the symptomatology of individuals with neurodevelopmental disorders suggest deficits change over the lifespan. A challenge for animal studies modeling these disorders is to effectively capture developmental trajectories, especially for cognitive abilities (44, 45). Ideally, this requires longitudinal behavioral testing to assess cognitive ability across development in the same animal. In this study, we demonstrate distinct developmental trajectories for different types of recognition memory in WT LEH rats using a battery of four spontaneous exploration-based tasks. The ability to exhibit nonassociative memory for objects and associative OCR memory is apparent by 4 to 6 weeks of age [consistent with previous findings (28)]. In contrast, we find that the ability to exhibit associative OPR and OPCR memory does not emerge until 7 weeks of age [but see (30)]. Whereas the Fmr1 KO rats showed intact memory at each time point in the early-developing OR and OCR tasks, the loss of FMRP specifically affected the later-developing cognitive abilities, with a delay in the OPR task, and inability to show memory in the OPCR task at any age. OPR and OPCR require the coordination of a number of intact brain circuits, including the prefrontal cortex (32–34). The observed abnormalities in OPR and OPCR are paralleled by an age-dependent deficit in LTP in the prefrontal cortex. Our finding that LTP is intact at 4 to 6 weeks, but impaired at 10 to 12 weeks of age in Fmr1 KO rats, suggests that the role of FMRP in mediating this form of plasticity changes over postnatal development. A similar conclusion can be drawn from our finding that mGluR-dependent LTD in CA1 is increased in Fmr1 KO rats at 1 month but not 2 months of age. Future experiments to determine whether the targets of FMRP change over this period or whether the mRNAs associated with polyribosomes differ (see below) (46) should be considered.
The characterization of developmental trajectories of cognitive function in rodent models of neurodevelopmental disorders is important because it provides a temporal framework for designing experiments to determine whether potential therapeutic interventions either prevent the emergence of deficits or reverse established deficits. On the basis of this framework, we treated animals with lovastatin starting at 4 weeks of age and showed that this intervention prevented the emergence of cognitive deficits in OPR and OPCR usually seen at 7 to 9 weeks in Fmr1 KO rats. Furthermore, even after treatment was terminated at 9 weeks, these cognitive abilities remained intact for at least another 3 months (the last time point tested). This is particularly important because untreated Fmr1 KO rats were unable to show OPCR at any age tested. Lovastatin also prevented the emergence of age-dependent deficits in FMRP-dependent synaptic plasticity in the prefrontal cortex of Fmr1 KO rats.
To investigate the mechanisms underlying the effect of lovastatin treatment, we next examined basal protein synthesis in the hippocampus. The increase in protein synthesis in rat hippocampus at 4 weeks of age (26) persisted until 6 months of age and was normalized by transient lovastatin treatment from 4 to 9 weeks. This extends previous findings from the mouse model of FXS showing that the acute lovastatin application to hippocampal slices corrects deficits in basal protein synthesis (17) by demonstrating that oral lovastatin administration can reverse this deficit once it has appeared. This reversal lasts for 4 months after removal of the lovastatin, suggesting that the transient inhibition of the ERK signaling pathway is sufficient to promote a long-term reset of protein synthesis in FXS. This effect appears to be inconsistent with the idea that increased protein synthesis in the hippocampus of Fmr1 KO rats is a direct consequence of the loss of FMRP binding to their target RNAs (18). An alternative explanation is that the increased basal protein synthesis observed in Fmr1 KO hippocampus actually reflects a compensatory response to the absence of FMRP, and this is no longer needed after appropriate treatment. Such an interpretation would suggest that most of the excess signal in the protein synthesis assay arises from translation of mRNAs that are not direct FMRP targets, which is supported by our recent study (46) using the translating ribosome affinity purification assay that detects mRNAs associated with ribosomes. We found that FMRP-targeted mRNAs were underrepresented in the ribosome-bound pool in Fmr1 KO CA1 hippocampal pyramidal neurons relative to WT controls (46). Furthermore, Chrm4 mRNA, which encodes the muscarinic acetylcholine receptor 4 (M4), showed an increase in ribosome association, and stimulation of M4 normalized many Fmr1 KO phenotypes. These findings suggest that the increase in translation in Fmr1 KO neurons is a compensatory response to the deletion of FMRP rather than a direct result, at least at the ages tested in these studies (46).
Treatment with lovastatin may remove the trigger for these compensatory changes by preventing the developmental emergence of the cellular or circuit dysfunction associated with the loss of FMRP. Cellular and circuit excitability dysregulation has been shown in a range of brain regions and cell types in Fmr1 KO mice (9, 10). Because circuit activity is known to regulate early neural development, lovastatin could be exerting its disease-modifying role through its regulation of mGluR-dependent neuronal excitability. This hypothesis is supported by our previous findings that acute lovastatin application rescues both audiogenic seizures and mGluR-dependent epileptiform activity in acute brain slices in Fmr1 KO mice (17). Furthermore, acute lovastatin treatment rescued the increase in mGluR-dependent LTD in CA1 in Fmr1 KO mice (17).
One potential limitation of this study is that although there was no alteration in weight gain between genotypes or treatment group, we have not been able to measure the dose of lovastatin received by each animal. Furthermore, we have not defined whether a critical period exists for the effectiveness of lovastatin treatment or defined an effective minimum treatment duration and we have focused on associative learning paradigms that rely on an animal's inherent attraction to novelty. Whether the beneficial effects of lovastatin can be generalized to other ages and other forms of learning is unknown, but we note that previous studies have reported this to be the case (17). Whether these findings will directly translate to clinical outcome for individuals with FXS is unknown. However, in this context, it is important to remember that our treatment regimen was initiated in 1-month-old rats. Although it is difficult to accurately estimate the corresponding age in humans, it is clear that this is much earlier than most individuals with FX enrolled in ongoing trials with lovastatin (NCT02680379, NCT02998151, and NCT02642653).
In summary, using assays of cognitive ability that rely on an animal's natural exploratory behavior, we have been able to demonstrate that early, brief treatment restores the normal developmental trajectory of associative memory acquisition that persists well into adulthood. This rescue is paralleled by physiological and biochemical rescue in the prefrontal cortex and hippocampus, respectively. The findings provide proof-of-concept evidence that FXS, and perhaps neurodevelopmental disorders more generally, may be amenable to transient, early intervention to permanently restore normal cognitive developmental trajectories.
The purpose of this study was to test the hypothesis that early therapeutic intervention can produce long-lasting benefits on cognition in a rat model of FXS. For behavioral studies, four groups of rats were used: WT control, WT treatment, KO control, and KO treatment; n = 12 per group based on power calculations on published data from the laboratory (32) (effect size = 1.34, α = 0.05, and power = 0.85). Littermates were housed in mixed-genotype cages (3 to 4 rats per cage), and cages were randomly assigned to control or treatment conditions [in line with the ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (47)]. For the protein synthesis experiments, sample sizes were chosen on the basis of power calculations on published data from the laboratory (17, 26, 42) (effect size = 2.12, α = 0.05, and power = 0.91); WT control, WT treatment, KO control, and KO treatment (n = 6 per group; at least two slices from each rat used to produce the value for each animal) were used for metabolic labeling. In the same fashion, for in vitro electrophysiology, sample sizes were chosen on the basis of power calculations on published data from the laboratory (17, 26, 42) (effect size = 1.45, α = 0.05, and power = 0.82); WT control (n = 9), WT treatment (n = 7), KO control (n = 9), and KO treatment (n = 8) (at least two slices from each rat used to produce the value for each animal) were used as labels. Experimenters were blind to the genotype and the treatment during data collection, scoring of behavioral data, and data analysis. For behavioral testing, we excluded data from trials in which animals show very low object exploration (less than 5 s of exploration per object or 15 s of total object exploration during the sample phase, or less than 15 s of total object exploration in the test phase). No effect of genotype was observed on the number of trials excluded because of insufficient exploration. Testing was always performed during the light phase of the cycle. All animal experiments were approved by the University of Edinburgh veterinary services before their start and were performed in accordance with the guidelines established by European Community Council Directive 2010/63/EU (22 September 2010) and by the Animal Care (Scientific Procedures) Act 1986.
Animals and treatment
Male LE-Fmr1em1/PWC, hereafter referred to as Fmr1 KO, and WT littermates, bred in-house and kept in a 12-hour/12-hour light/dark cycle with ad libitum access to water and food, were used. Colony founders were produced by SAGE (Sigma Advanced Genetic Engineering) Labs using ZFN-mediated disruption of Fmr1 (48) with a targeted construct containing coding sequence for eGFP; resulting founders did not express FMRP or eGFP. Pups were weaned off their dams at P22 and housed in mixed-genotype cages with littermates (three to four animals per cage). Animals were genotyped by PCR. Ad libitum standard laboratory chow was provided until P29. On P29, the diet was changed to either control or lovastatin-enriched (100 mg/kg; Bio-Serv) diet, which was restocked and weighed once daily. At P64, animals were returned to ad libitum standard laboratory chow until the end of experiment (P164). Rats' weight and consumption per cage were monitored throughout the dosing period (P29 to P64) to ensure that their diet did not have any adverse effects on their growth.
RNA isolation and RT-PCR
Total hippocampal RNA was isolated from 4-month-old rats (three WT and three Fmr1 KO) using the RNeasy Lipid Tissue Kit (Qiagen) as per the manufacturer's instructions. Two micrograms of total RNA was used for cDNA synthesis using SuperScript III (Invitrogen) with oligo(dT) and random hexamers. PCR was performed using the GoTaq Green Master Mix (Promega). Fmr1 primers span exons 1 to 4 [Fmr1_e1F: CGA GGA AGG ACG AGA AGA TG and Fmr1_e4R: CAC CCT TTA TCA TCC TCA C; amplicon, 284 base pair (bp)]. Primers to GFP (GFP_F: ACG TAA ACG GCC ACA AGT TC and GFP_R: ATG CCG TTC TTC TGC TTG TC; amplicon, 421 bp) and 18S (18S_F: GTG GAG CGA TTT GTC TGG TT and 18S_R: CAA GCT TAT GAC CCG CAC TT; amplicon, 321 bp) and cDNA from a GFP transgenic mouse were used as positive controls.
Hippocampal extracts from Fmr1 KO rats and controls (n = 3 per age for developmental expression; n = 3 per genotype at P14 to verify the loss of expression) were prepared in radioimmunoprecipitation assay buffer containing protease inhibitors (cOmplete EDTA-free), immunoblotted using primary antibodies raised to the C-terminal half of FMRP (1:5000; ab69815, AbCAM), GFP (1:5000; ab6673, AbCAM), and β-actin (1:10,000; AC-74, Sigma-Aldrich) and were imaged as previously described in (36).
Histology was performed as previously described (49). Coronal sections were reacted with an antibody raised to the N-terminal half of FMRP (1:1500; MAB2160, Millipore).
Spontaneous object exploration tasks
Apparatus. Animals were tested in a rectangular polycarbonate testing box (76 cm long by 45 cm wide by 60 cm tall) with removable walls and floor inserts that could conform to two contexts. In context 1, white textured wallpaper and wood-effect linoleum floor were used. Two 3M Dual-Lock resealable fasteners were attached to the floor, 9 cm from the box walls at north-east and north-west locations, used to keep the two objects firmly attached to the floor in the same locations for every trial. In context 2, matte blue–painted walls and black rubber-textured floor insert were used, with holes cut to gain access to the Dual-Lock resealable fasteners where the objects were attached. The testing box was placed on a table surrounded on three sides by a black curtain, with one opening at the south side of the box (where subjects were placed). A lamp in the north-west corner and a large high contrast poster on the north-east corner were used as external cues, which remained in the same position and orientation throughout the experiments. The external environment was kept as consistent as possible, and a radio on low volume was used to mask potentially distracting noises. A variety of objects were used, which were between 8 cm by 8 cm by 8 cm and 11 cm by 11 cm by 11 cm, were nonporous, and were easily cleaned. Each object was used only once per animal. Objects were cleaned between trials with 70% ethanol solution and unscented baby wipes (Huggies).
Handling and habituation. Starting from P21 (a day before weaning), animals were handled daily in the animal house and experimental room for 7 days before experiments. Task-specific habituation was performed 2 days before experiments (P26 to P27) within the experimental apparatus box to familiarize the animals to the apparatus and type of objects. On P26, the animals were habituated to both context configurations in cage groups (30 min per context) in the morning and individually (10 min per context) in the afternoon. Between exposures to contexts, rats were placed in an opaque holding bucket. On P27, animals were individually habituated twice (morning and afternoon both context configurations each time), but this time, two different objects were fixed in the positions where they would encounter objects during testing (10 min per context, with objects). These objects were not used again during testing. During the habituation sessions, subjects were left undisturbed to explore.
Testing: General procedures. Rats were tested on four different object exploration tasks (OR, OCR, OPR, and OPCR) over a 2-day period, and testing was repeated in the same rats at different ages (P28 and P29, P35 and P36, P42 and P43, P49 and P50, P56 and P57, P63 and P64, P70 and P71, P77 and P78, P98 to P105, and P164 and P165). The general procedures for each task were the same.
For each phase of every task, the experimenter prepared the appropriate context configuration and attached two cleaned appropriate objects to the appropriate locations within the box using the Dual-Lock resealable fasteners. Each animal was then removed from the home cage and placed in the box, facing the south wall of the apparatus. The sample phases and test phases were each 3 min long, during which the animals were free to explore. An overhead black and white camera (Panasonic) was used to monitor the exploring rat around the testing box. The video signal was fed into a DVD recorder and a computer on the desk of the experimenter, which was 2 m away from the testing box. The computer ran an in-house timing software (National Instruments, LabVIEW), whereby the press of a key on the computer mouse would activate a timer. This was performed manually by the experimenter who observed the behavior of the rat via the computer screen and recorded the amount of time that the rat was engaged in exploration of each object. After 3 min, the animal was either placed in the opaque holding bucket (30 cm in diameter, containing standard bedding) for 2 min (after a sample phase) or returned to the home cage (after a test phase). Exploration was defined as the animal actively exploring an object with its snout within 2 cm of the object and performing actions such as sniffing and whisking. Exploration was not scored when the animal was not actively exploring object (i.e., climbing or resting on an object). Novel object positions, test phase contexts, context order in OCR and OPCR, and objects were counterbalanced across genotypes, tasks, and time points to ensure that the final results were as unbiased as possible.
Object recognition. OR (Fig. 1A) is a two-phase nonassociative recognition task. In the sample phase, two identical objects are available in either context 1 or context 2. In the test phase, two objects (one identical to the objects from the sample phase and one novel object) are available in the same context as the sample phase. This task is used to test whether the animal can detect object novelty and discriminate between two nonidentical objects. Higher exploration of the novel than the familiar object is indicative of memory for the familiar object.
Object-place recognition. OPR (Fig. 1A) is a two-phase associative recognition task. In the sample phase, two nonidentical objects are available in either context 1 or context 2. In the test phase, two objects (both identical to one object from the sample phase) are available in the same context as in the sample phase. This task is used to test whether an animal can associate a specific object with a location in space. Higher exploration of the object that is in a different location than it was experienced in the sample phase is indicative of OPR memory.
Object-context recognition. OCR (Fig. 1A) is a three-phase associative recognition task. In sample phase 1, two identical objects are available in either context 1 or context 2. In sample phase 2, a different pair of identical objects is explored in the other context. In the test phase, two objects (one identical to the objects from sample phase 1 and the other identical to the objects from sample phase 2) are available in either context 1 or context 2. This task is used to test whether an animal can associate an object with a surrounding context. Higher exploration of the object, which is in a different context than it was experienced in the sample phase, is indicative of OCR memory.
Object-place-context recognition. OPCR (Fig. 1A) is a three-phase associative recognition task. In sample phase 1, two nonidentical objects are available in either context 1 or context 2. In sample phase 2, objects identical to those in sample phase 1 are available, but the objects have swapped locations and are in the other context. In the test phase, two identical objects (identical to one of the objects in sample phases 1 and 2) are available in one of the two contexts. This task is used to test whether the animal can associate an object to a location in a specific surrounding context (episodic-like memory). Higher exploration of the object, which is in a different location in that context than it was in the sample phase, is indicative of OPCR memory.
In vitro electrophysiology
Medial prefrontal cortex long-term potentiation. Subjects were anesthetized using isoflurane and decapitated. The brain was quickly dissected out and placed in ice-cold (<4°C) modified slicing artificial cerebrospinal fluid (ACSF) solution (NaCl, 86 mM; NaH2PO4, 1.2 mM; KCl, 2.5 mM; NaHCO3, 25 mM; d-glucose, 24 mM; sucrose, 75 mM; CaCl2, 0.5 mM; and MgCl2, 7 mM). Coronal slices (300 μm) containing prelimbic mPFC were cut in ice-cold modified slicing ACSF solution using a vibratome and transferred to a holding chamber containing warmed recording ACSF solution (NaCl, 124 mM; NaH2PO4, 1.2 mM; KCl, 2.5 mM; NaHCO3, 25 mM; d-glucose, 20 mM; CaCl2, 2 mM; and MgCl2, 1 mM), where they were maintained at 35°C for 30 min. Slices were left to recover for a further 30 min at room temperature before the start of experimentation.
Slices were then placed in a submerged recording chamber heated to 31°C and perfused with prewarmed carbogenated recording ACSF solution at a rate of 4 to 5 ml/min. A recording electrode was placed in layer 5 of the prelimbic mPFC, and stimulating electrode was placed in either layer 2/layer 3. Stimulating and recording electrodes were staggered to prevent direct antidromic stimulation. Synaptic responses were evoked every 30 s using a bipolar nichrome stimulating wire attached to a constant current stimulus isolator, delivering a 200-μs pulse. After the acquisition of a 20-min stable baseline, LTP was induced using 5× 500-ms trains of 300-Hz stimulation at 3-min intervals. Responses were then recorded for 60 min after tetanization.
Signal waveforms were amplified 1000×, low pass–filtered at 4 kHz, and digitized at 20 kHz. fEPSP slopes were normalized to baseline values. Magnitude of LTP was calculated from 40 to 60 min after tetanus time points. Normalized data were averaged across experimental groups and reported as means ± SE.
DHPG-induced LTD. Horizontal hippocampal slices (400 μm) prepared from P21 to P32 animals were incubated in oxygenated ACSF at 31°C for 30 min and then stored at room temperature until recording. An incision was made through CA3 before recording. Slices were continuously perfused in an interface chamber with 30° ± 1°C ACSF saturated with 95% O2 and 5% CO2 at 4 to 5 ml/min. mGluR-LTD was induced using S-DHPG (50 μM) for 5 min. LTD magnitude was calculated by dividing the average fEPSP slope from 50 to 60 min after DHPG application by the average fEPSP slope during the 20-min baseline before DHPG application.
Metabolic labeling. Hippocampal slices were prepared from age-matched male WT and Fmr1 KO rats in an interleaved fashion as previously described (17, 26, 42). Briefly, hippocampi were rapidly isolated, and 500-μm slices were prepared from the dorsal half using a Stoelting tissue slicer. Slices were recovered for 4 hours in 32.5°C ACSF (NaCl, 124 mM; KCl, 3 mM; NaH2PO4, 1.25 mM; NaHCO3, 26 mM; dextrose, 10 mM; MgCl2, 1 mM; and CaCl2, 2 mM, saturated with 95% O2 and 5% CO2) and then incubated for 30 min with 25 μM ActD to block transcription. To measure protein synthesis, slices were then transferred to fresh ACSF containing 35S-Met/Cys (10 μCi/ml; PerkinElmer) and incubated for 30 min. After labeling, slices were homogenized, and labeled proteins were isolated by trichloroacetic acid precipitation. Samples were read with a scintillation counter and subjected to a protein concentration assay (Bio-Rad). Final data were expressed as counts per minute per microgram of protein, normalized to the 35S-Met/Cys ACSF used for incubation, and the average incorporation of all samples was analyzed in that experiment (eight samples per experiment: four WT and four Fmr1 KO). All aspects of the experiments were performed blinded to genotype.
For all experiments, the researchers conducting the experiments and scoring and analyzing the data were blinded to the genotype of the rats and to treatment group. For the lovastatin dosing experiments, mixed-genotype cages were randomly allocated to lovastatin and control diet groups. Statistical analysis was performed using GraphPad (Prism 6.0), SPSS 16.0 software, or R v.3.4.4 (R Core Team, 2018); scripts were written and run using RStudio 1.0.153 (RStudio Team, 2016). Because of missing data due to our exclusion criteria, LMEs were fitted to our longitudinal behavioral data using the R package lme4 v1.1-17 (50). Animal identity was included in models as a random effect, and the variables of interest were included as fixed effects. To evaluate the significance of effects using LMEs, the model without the variable of interest (a reduced/null model) was compared to the model with the variable of interest using a likelihood ratio test, where the appropriate statistical significance was assessed using either two- or three-way ANOVA. Unpaired two-sample t tests followed by Bonferroni correction were used to compare differences between groups. One-sample t tests were used to compare DIs against chance (DI = 0) controlled for the false discovery rate using the Benjamini-Hochberg procedure (51). Rats were used as an experimental unit throughout the manuscript. In electrophysiological experiments, where multiple slices were used from each animal (LTP/LTD experiments, protein synthesis), the average value for each animal was used in the analysis. Results are presented as means ± SEM. Probabilities of P < 0.05 were considered as significant. The raw data are provided in data file S1.
stm.sciencemag.org/cgi/content/full/11/494/eaao0498/DC1
Fig. S1. Adult Fmr1 KO LEH rats exhibit deficits in OPCR.
Fig. S2. Object exploration time during the test phase of spontaneous recognition tasks does not differ between genotypes.
Fig. S3. Rationale of lovastatin treatment in FXS.
Fig. S4. Lovastatin has negligible effect on the weight gain of rats.
Fig. S5. Object exploration time during the test phase of spontaneous recognition tasks does not differ between genotypes with or without lovastatin treatment.
Fig. S6. WT and Fmr1 KO rats perform equally well in OR and OCR tasks throughout development with control and lovastatin diets.
Fig. S7. Fmr1 KO LEH rats exhibit age-specific increase of group I mGluR-LTD in CA1 of the hippocampus.
Table S1. Statistical results from one-sample t tests and post hoc two-sample t tests for object exploration tasks throughout development in WT and Fmr1 KO rats.
Table S2. Statistical results from two-way ANOVA of exploration times in object exploration tasks throughout development in WT and Fmr1 KO rats.
Table S3. Statistical results from one-sample t tests for object exploration tasks throughout development in WT and Fmr1 KO rats with or without lovastatin treatment.
Table S4. Statistical results from post hoc two-sample t tests for object exploration tasks throughout development in WT and Fmr1 KO rats with or without lovastatin treatment.
Table S5. Statistical results from two-way ANOVA of exploration times in object exploration tasks throughout development in WT and Fmr1 KO rats with or without lovastatin treatment.
Table S6. Statistical results from adult object exploration tasks, effect of lovastatin on food intake/weight gain, hippocampal basal protein synthesis, and synaptic plasticity data.
Table S7. LME model distribution tests of behavioral data.
Table S8. LME modeling results of WT and Fmr1 KO object exploration tasks throughout development.
Table S9. LME modeling results of WT and Fmr1 KO object exploration tasks throughout development with or without lovastatin treatment.
Data file S1. Raw data (provided as separate Excel file).
W. E. Kaufmann,
S. A. Kidd,
H. F. Andrews,
D. B. Budimirovic,
A. Esler,
B. Haas-Givler,
T. Stackhouse,
C. Riley,
G. Peacock,
S. L. Sherman,
W. T. Brown,
E. Berry-Kravis
, Autism spectrum disorder in fragile X syndrome: Cooccurring conditions and current treatment. Pediatrics 139, S194–S206 (2017).
R. J. Hagerman,
E. Berry-Kravis,
D. B. Bailey Jr.,
H. Moine,
R. F. Kooy,
F. Tassone,
I. Gantois,
N. Sonenberg,
J. L. Mandel,
P. J. Hagerman
, Fragile X syndrome. Nat. Rev. Dis. Primers 3, 17065 (2017).
E. G. Harlow,
S. M. Till,
T. A. Russell,
L. S. Wijetunge,
P. Kind,
A. Contractor
, Critical period plasticity is disrupted in the barrel cortex of Fmr1 knockout mice. Neuron 65, 385–398 (2010).
Q. He,
T. Nomura,
J. Xu,
, The developmental switch in GABA polarity is delayed in fragile X mice. J. Neurosci. 34, 446–450 (2014).
A. J. M. H. Verkerk,
M. Pieretti,
J. S. Sutcliffe,
Y.-H. Fu,
D. P. A. Kuhl,
A. Pizzuti,
O. Reiner,
S. Richards,
M. F. Victoria,
F. Zhang,
B. E. Eussen,
G.-J. B. van Ommen,
L. A. J. Blonden,
G. J. Riggins,
J. L. Chastain,
C. B. Kunst,
H. Galjaard,
C. Thomas Caskey,
D. L. Nelson,
B. A. Oostra,
S. T. Warran
, Identification of a gene (FMR-1) containing a CGG repeat coincident with a breakpoint cluster region exhibiting length variation in fragile X syndrome. Cell 65, 905–914 (1991).
K. De Boulle,
E. Reyniers,
L. Vits,
J. Hendrickx,
B. Van Roy,
F. Van Den Bos,
E. de Graaff,
P. J. Willems
, A point mutation in the FMR-1 gene associated with fragile X mental retardation. Nat. Genet. 3, 31–35 (1993).
J. B. Zang,
E. D. Nosyreva,
C. M. Spencer,
L. J. Volk,
K. Musunuru,
R. Zhong,
E. F. Stone,
L. A. Yuva-Paylor,
K. M. Huber,
R. Paylor,
J. C. Darnell,
R. B. Darnell
, A mouse model of the human fragile X syndrome I304N mutation. PLOS Genet. 5, e1000758 (2009).
M. S. Sidorov,
B. D. Auerbach,
M. F. Bear
, Fragile X mental retardation protein and synaptic plasticity. Mol. Brain 6, 15 (2013).
A. Contractor,
V. A. Klyachko,
C. Portera-Cailliau
, Altered neuronal and circuit excitability in fragile X syndrome. Neuron 87, 699–715 (2015).
S. Chattarji,
D. J. A. Wyllie,
P. C. Kind
, Fragile X syndrome: From targets to treatments. Neuropharmacology 68, 83–96 (2013).
J. F. Werker,
T. K. Hensch
, Critical periods in speech perception: New directions. Annu. Rev. Psychol. 66, 173–196 (2015).
F. Sengpiel,
, The role of activity in development of the visual system. Curr. Biol. 12, R818–R826 (2002).
R. M. Meredith
, Sensitive and critical periods during neurotypical and aberrant neurodevelopment: A framework for neurodevelopmental disorders. Neurosci. Biobehav. Rev. 50, 180–188 (2015).
E. M. Berry-Kravis,
L. Lindemann,
A. E. Jønch,
G. Apostol,
M. F. Bear,
R. L. Carpenter,
J. N. Crawley,
A. Curie,
V. Des Portes,
F. Hossain,
F. Gasparini,
B. Gomez-Mancilla,
D. Hessl,
S. H. Scharf,
P. P. Wang,
F. Von Raison,
R. Hagerman,
W. Spooren,
S. Jacquemont
, Drug development for neurodevelopmental disorders: Lessons learned from fragile X syndrome. Nat. Rev. Drug Discov. 17, 280–299 (2018).
M. R. Tranfaglia,
C. Thibodeaux,
D. J. Mason,
I. Roberts,
R. Smith,
T. Guilliams,
P. Cogram
, Repurposing available drugs for neurodevelopmental disorders: The fragile X experience. Neuropharmacology 147, 74–86 (2019).
C. Henderson,
L. Wijetunge,
M. N. Kinoshita,
M. Shumway,
R. S. Hammond,
F. R. Postma,
C. Brynczka,
R. Rush,
A. Thomas,
S. T. Warren,
P. W. Vanderklish,
P. C. Kind,
A. M. Healy
, Reversal of disease-related pathologies in the fragile X mouse model by selective activation of GABAB receptors with arbaclofen. Sci. Transl. Med. 19, 152ra128 (2012).
E. K. Osterweil,
S.-C. Chuang,
A. A. Chubykin,
M. Sidorov,
R. Bianchi,
R. K. S. Wong,
, Lovastatin corrects excess protein synthesis and prevents epileptogenesis in a mouse model of fragile X syndrome. Neuron 77, 243–250 (2013).
S. T. Warren
, The mGluR theory of fragile X mental retardation. Trends Neurosci. 27, 370–377 (2004).
J. Visootsak,
D. Budimirovic,
M. Cherubini,
P. Zarevics,
K. Walton-Bowen,
P. Wang,
R. L. Carpenter
, Arbaclofen in fragile X syndrome: Results of phase 3 trials. J. Neurodev. Disord. 9, 3 (2017).
A. Çaku,
D. Pellerin,
P. Bouvier,
E. Riou,
F. Corbin
, Effect of lovastatin on behavior in children and adults with fragile X syndrome: An open-label study. Am. J. Med. Genet. A 164, 2834–2842 (2014).
D. E. Berlyne
, Novelty and curiosity as determinants of exploratory behaviour. Br. J. Psychol. Gen. Sect. 41, 68–80 (1950).
A. Ennaceur,
J. Delacour
, A new one-trial test for neurobiological studies of memory in rats. 1: Behavioral data. Behav. Brain Res. 31, 47–59 (1988).
S. L. Dix,
J. P. Aggleton
, Extending the spontaneous preference test of recognition: Evidence of object-location and object-context recognition. Behav. Brain Res. 99, 191–200 (1999).
G. Norman,
M. J. Eacott
, Dissociable effects of lesions to the perirhinal cortex and the postrhinal cortex on memory for context and objects in rats. Behav. Neurosci. 119, 557–566 (2005).
M. J. Eacott,
G. Norman
, Integrated memory for object, place, and context in rats: A possible model of episodic-like memory? J. Neurosci. 24, 1948–1953 (2004).
A. Asiminas,
A. D. Jackson,
D. Katsanevaki,
S. A. Barnes,
E. R. Wood,
, Conserved hippocampal cellular pathophysiology but distinct behavioural deficits in a new rat model of FXS. Hum. Mol. Genet. 24, 5977–5984 (2015).
S. R. Westbrook,
L. E. Brennan,
M. E. Stanton
, Ontogeny of object versus location recognition in the rat: Acquisition and retention effects. Dev. Psychobiol. 56, 1492–1506 (2014).
A. I. Ramsaran,
, Ontogeny of object-in-context recognition in the rat. Behav. Brain Res. 298, 37–47 (2016).
J. A. Ainge,
R. F. Langston
, Ontogeny of neural circuits underlying spatial memory in the rat. Front. Neural Circuits 6, 8 (2012).
H. R. Sanders,
, Determinants of object-in-context and object-place-context recognition in the developing rat. Dev. Psychobiol. 58, 883–895 (2016).
M. W. Brown,
, Recognition memory: What are the roles of the perirhinal cortex and hippocampus? Nat. Rev. Neurosci. 2, 51–61 (2001).
R. F. Langston,
E. R. Wood
, Associative recognition and the hippocampus: Differential effects of hippocampal lesions on object-place, object-context and object-place-context memory. Hippocampus 20, 1139–1153 (2010).
O. Y. Chao,
J. P. Huston,
J.-S. Li,
A.-L. Wang,
M. A. de Souza Silva
, The medial prefrontal cortex—Lateral entorhinal cortex circuit is essential for episodic-like memory and associative object-recognition. Hippocampus 26, 633–645 (2016).
D. I. G. Wilson,
S. Watanabe,
H. Milner,
J. A. Ainge
, Lateral entorhinal cortex is necessary for associative but not nonassociative recognition memory. Hippocampus 23, 1280–1290 (2013).
M. I. Schlesiger,
M. Wagner,
, Lateral entorhinal cortex is critical for novel object-context recognition. Hippocampus 23, 352–366 (2013).
C. Bostrom,
S. -y. Yau,
N. Majaess,
M. Vetrici,
J. Gil-Mohapel,
B. R. Christie
, Hippocampal dysfunction and cognitive impairment in Fragile-X Syndrome. Neurosci. Biobehav. Rev. 68, 563–574 (2016).
S. R. Hooper,
D. Hatton,
J. Sideris,
K. Sullivan,
P. A. Ornstein,
D. B. Bailey Jr..
, Developmental trajectories of executive functions in young males with fragile X syndrome. Res. Dev. Disabil. 81, 73–88 (2018).
A. Michalon,
T. M. Ballard,
L. Ozmen,
J. G. Wettstein,
G. Jaeschke,
L. Lindemann
, Chronic pharmacological mGlu5 inhibition corrects fragile X in adult mice. Neuron 74, 49–56 (2012).
M. Lambert,
P.-J. Lupien,
C. Gagné,
E. Lévy,
S. Blaichman,
S. Langlois,
M. Hayden,
V. Rose,
J. T. R. Clarke,
B. M. J. Wolfe,
C. Clarson,
H. Parsons,
D. K. Stephure,
D. Potvin,
J. Lambert, Canadian Lovastatin in Children Study Group
, Treatment of familial hypercholesterolemia in children and adolescents: Effect of lovastatin. Pediatrics 97, 619–628 (1996).
G. Dölen,
E. Osterweil,
B. S. S. Rao,
G. B. Smith,
, Correction of fragile X syndrome in mice. Neuron 56, 955–962 (2007).
D. D. Krueger,
K. Reinhold,
, Hypersensitivity to mGluR5 and ERK1/2 leads to excessive protein synthesis in the hippocampus of a mouse model of fragile X syndrome. J. Neurosci. 30, 15616–15627 (2010).
N. H. Komiyama,
S. G. N. Grant,
U. V. Nägerl,
D. J. A. Wyllie
, Convergence of hippocampal pathophysiology in Syngap+/− and Fmr1−/y mice. J. Neurosci. 35, 15073–15081 (2015).
H. G. S. Martin,
O. Lassalle,
J. T. Brown,
O. J. Manzoni
, Age-dependent long-term potentiation deficits in the prefrontal cortex of the Fmr1 knockout mouse model of fragile X syndrome. Cereb. Cortex 26, 2084–2092 (2016).
J. K. Y. Lai,
M. Sobala-Drozdowski,
L. Zhou,
L. C. Doering,
P. A. Faure,
J. A. Foster
, Temporal and spectral differences in the ultrasonic vocalizations of fragile X knock out mice during postnatal development. Behav. Brain Res. 259, 119–130 (2014).
S.-W. Yun,
J. Platholi,
M. S. Flaherty,
W. Fu,
A. H. Kottmann,
M. Toth
, Fmrp is required for the establishment of the startle response during the critical period of auditory development. Brain Res. 1110, 159–165 (2006).
S. R. Thomson,
S. S. Seo,
S. R. Louros,
M. Muscas,
O. Dando,
C. Kirby,
G. E. Hardingham,
E. K. Osterweil
, Cell-type-specific translation profiling reveals a novel strategy for treating fragile X syndrome. Neuron 95, 550–563.e5 (2017).
C. Kilkenny,
W. Browne,
I. C. Cuthill,
M. Emerson,
D. G. Altman
, Animal research: Reporting in vivo experiments: The ARRIVE guidelines. Br. J. Pharmacol. 160, 1577–1579 (2010).
A. M. Geurts,
G. J. Cost,
Y. Freyvert,
B. Zeitler,
J. C. Miller,
V. M. Choi,
S. S. Jenkins,
A. Wood,
X. Cui,
X. Meng,
A. Vincent,
M. Michalkiewicz,
R. Schilling,
J. Foeckler,
S. Kalloway,
H. Weiler,
S. Ménoret,
I. Anegon,
G. D. Davis,
E. J. Rebar,
P. D. Gregory,
F. D. Urnov,
H. J. Jacob,
R. Buelow
, Knockout rats via embryo microinjection of zinc-finger nucleases. Science 325, 433 (2009).
V. G. Seidel,
E. Harlow,
A. K. Wright,
C. Bagni,
T. H. Gillingwater,
, Altered maturation of the primary somatosensory cortex in a mouse model of fragile X syndrome. Hum. Mol. Genet. 21, 2143–2156 (2012).
D. Bates,
M. Mächler,
B. Bolker,
S. Walker
, Fitting linear mixed-effects models using lme4. J. Stat. Softw. 67, 1–48 (2015).
Y. Benjamini,
Y. Hochberg
, Controlling the false discovery rate: A practical and powerful approach to multiple testing. J. R. Stat. Soc. Series B 57, 289–300 (1995).
Acknowledgments: We would like to thank A. Aruldass, K. Reed, and C. Neill-Edwards for assistance in behavioral data collection and colleagues in the Wood and Kind groups and the Simons Initiative for the Developing Brain for constructive discussions during the course of this study. Funding: A.A. was the recipient of a fellowship from the Greek State Scholarship Foundation IKY (Maria Zaousi bequest). This study was supported by grants from SFARI, The Shirley Foundation, Medical Research Council UK (MR/P006213/1), FRAXA Research Foundation, Wadhwani Foundation, The Patrick Wild Centre, Indian Department of Biotechnology, and RS Macdonald Charitable Trust. This study was also supported by the Wellcome Trust/Royal Society (Sir Henry Dale fellowship 104116/Z/14/Z) and Medical Research Council (MRC MR/M006336/1). This research was also supported, in part, by grants to M.F.B. from NIMH (R01MH106469) and the Simons Foundation (SFARI #240559). Author contributions: A.A., A.D.J., S.M.T., S.R.L., E.R.W., and P.C.K. conceived and designed the experiments. A.A. performed the behavioral experiments and analyzed the results. A.D.J. performed the electrophysiology experiments and analyzed the results. S.M.T. performed the immunohistochemistry and molecular biology experiments and analyzed the results. S.R.L. performed the biochemistry experiments and analyzed the results. T.S. performed the electrophysiology experiments. O.D. performed the statistical analysis. S.M.T. and P.C.K. designed the genetically modified rats. M.F.B., D.J.A.W., G.E.H., S.C., and E.K.O. helped guide the research. A.A., A.D.J., S.M.T., E.R.W., and P.C.K. interpreted all the results and wrote the paper. Competing interests: M.F.B. holds a patent titled "Methods of treating disorders with group I mGluR antagonists" (US6890931B2) and has served as a paid consultant to Q-State Biosciences, Vertex Pharmaceuticals, and Sunovion Pharmaceuticals. Data and materials availability: The LE-Fmr1em1/PWC rat line is available from P.C.K. under a material transfer agreement with the University of Edinburgh. All data associated with this study are present in the paper or Supplementary Materials.
Vol 11, Issue 494
You are going to email the following Sustained correction of associative learning deficits after brief, early treatment in a rat model of Fragile X Syndrome
By Antonis Asiminas, Adam D. Jackson, Susana R. Louros, Sally M. Till, Teresa Spano, Owen Dando, Mark F. Bear, Sumantra Chattarji, Giles E. Hardingham, Emily K. Osterweil, David J. A. Wyllie, Emma R. Wood, Peter C. Kind
Science Translational Medicine 29 May 2019
Brief, early treatments with lovastatin provide long-lasting rescue of associative memory deficits in a rat model of Fragile X.

redpajama_set_name: RedPajamaCommonCrawl


Although less well represented onscreen the vast majority of the Imperial war machine consists of basic infantry. Generally less well trained than the elite Stormtrooper, the grey clad infantry man is on many worlds the primary weapons of the empire.
First seen in Return of the Jedi, then given a whole new spin in Rogue One, Rebel Commandos are the elite ground foces of the rebellion.
A long time coming but I managed to finish and base four squads of Rebel Troopers. Sculpted by Bob Charrette, Julie Guthrie & Johnathan Woods. Lovely minis with a delicate touch on the fabric, very different from the Stormtroopers.
With my newly painted stormtroopers ready for action it was time to dust off the Star Wars Miniatures Battles rules. The last time these had seen action was in the mid nineties so a revision of the rules was in order.
Like most figure painters I did not relish the task of painting the trooper's armour.
Perhaps no other sci-fi goon is as recognisable as the faceless minion of evil than the Imperial Stormtrooper. The elite of the Empire whose death's head inspired helmet and stark white armour represent the cold faceless power of totalitarian rule.
This miniature seems to perfectly capture the original D&D cleric, there is even a hint of Easley in the sculpt! The Adventuring Cleric, No. 891 from the Personalities range.
Sold as Female Elf Ranger, No. 885 from the Personalities range. I felt plate armour was too much for rangers so instead went for a human female knight. Her cloak was a bit of a colour experiment and I was happy with the result.
Sold as Knight with Lion Shield No. 835 from the Personalities range. A classic paladin!


Metal punching processes are widely used in mass production because of their simplicity, high levels of productivity, and low costs. The quality of the product made by punching greatly depends on various processing parameters such as the tool geometries, sheet material properties, punching conditions, punch and die material properties, and tooling wears. In this study, the effects of die clearance angles and die opening sizes were compared on the accuracy and quality of punched holes produced on 1.4-mm thick S275 mild steel metal sheet. A total of nine punching dies were made with die clearance angles of 0.25°, 0.50°, and 0.75° for each die opening sizes of ø20.225 mm, ø13.225 mm, and ø12.225 mm. The burr formations and die weight losses were monitored for 10th, 50th, 100th, 500th, and 1000th strokes. The results show that the die weight loss, indicative of wear, increases as the number of strokes increase, with the most losses at the initial stages of the punching process (10th–50th strokes). The lowest weight loss was obtained for dies with 0.75° angular clearance, corresponding to the lowest wear occurrence. The low wear indicates that the shearing edge remains relatively sharp and thus able to produce good quality holes. Thus, for producing good quality holes and increasing tool life, it is recommended that an angular clearance of 0.75° is considered in the design of punching dies.
The authors express their gratitude to the Department of Mechanical and Manufacturing, Advanced Technology Training Center (ADTEC) Kulim, Kedah, for providing the use of heat treatment, hardness testing and WEDM facilities and the School of Mechanical Engineering, Universiti Sains Malaysia, Penang, for providing access to press machines and SEM facilities.
The first author would like to acknowledge the Public Service Department of Malaysia for the scholarship support under the HLP program.


State Congress just closed, and Texas veterans are walking away with a number of new programs and benefits.
As of Sept. 1, veterans and their spouses will be waived of application and examination fees for occupational licenses. A new grant program will support community-based mental health programs for veterans and their families with a $10,000,000 per year price tag. A pilot preventative services program will be established for vets considered to be at high risk of domestic violence as well as both a rural and women's mental health initiative and increasing accessibility to counseling- both peer-to-peer and with licensed professionals.
The most notable bill, HB 867, established the first permanent state program for women veterans in Texas -The Texas Women's Veterans Program.
The program has been functioning under The Texas Veterans Commission as the Women Veterans Initiative – started in 2012 with four employees. Advocates have since fought to make the program permanent.
The bill faced opposition – the Commission was not convinced of the necessity for a specific program serving women vets. Upon hearing testimony from women veterans in House and Senate committees, that emphasized their higher risk of sexual assault, the need for prenatal care and the fact that the program would run on existing funds and grants – the bill passed.
According to the U.S. Department of Veterans Affairs, one in four veteran women have experienced military sexual trauma – exclusively female spaces help women feel secure in a safe environment. Female spaces can better cater to the needs of veterans who are also single mothers and often need as much help with childcare and schooling as well as assistance with employment and housing. When women return from the service, they do not face different challenges, but potentially face more. The permanent title will allow the program to expand while increasing grant and donation funding.
"The program will continue to connect women with the benefits and services they have earned," said Edith Disler, manager of the Women Veterans Initiative.
Through conferences, seminars and training workshops the program will provide guidance and direction to women vets who apply for grants, benefits or services, as well as perform outreach functions to inform these vets of their eligibility.
Disler also hopes to work with local entities to target the needs of women student vets.
"It's not that I've felt isolated, it's just I haven't always had help," junior business major Bethany Grant said. "If this was already out there, I missed it. I mean, I know basically what's out there, but I had a hard time when the semester started, working out daycare. I'll definitely be checking in next semester," she continued.


text:
Washington, March 10, 2014 – The National Council of Churches first "Christian Unity Gathering" of leaders from member communions and other partners will be held May 18-20, 2014, in the Hilton Washington, Dulles, Herndon, Va.
"We are getting together to celebrate ecumenism and energize our journey toward the visible unity of all the Christian churches," said Jim Winkler, President and General Secretary of the NCC.
The Rev. Ann Tiemeyer, NCC Interim Associate General Secretary for Joint Action and Advocacy, said the program "will include ample opportunity for all attendees to engage with one another through group discussion, input from expert resource people to help inform our discussion, and celebratory service with words from interfaith guests and an address from Jim Winkler."
Addressing the Gathering on the NCC priority of working against mass incarceration will be Dr. Iva E. Carruthers, social justice and policy advocate, educator and activist, and the general secretary of the Samuel DeWitt Proctor Conference; Dr. Marian Wright Edelman, founder and president of the Children's Defense Fund; the Rev. Dr. Harold Dean Trulear, Associate Professor of Applied Theology and Director of the Doctor of Ministry program, Howard University; and Jim Wallis, founder and president of Sojourners, and editor of Sojourners magazine.
This is the NCC's first national gathering since it reorganized last year, reduced the size of the Governing Board, and created four 'convening tables' responsible for key mission and programs.
The Christian Unity Gathering will be the first national event to convene the tables. The main focus of their work will be interreligious relations with a focus on peace and mass incarceration.
The Council's response to mass incarceration will be fleshed out by the May Gathering, and by the Council's Governing Board.
Likely to be included in the discussions are the realities that the U.S. has the largest prison population in the world, that there is an enormous racial and ethnic imbalance both in the prison population and on death row, that there are many persons imprisoned whose crimes are victimless, non-violent, or related to personal drug possession and use, and that the billions of dollars spent keeping people locked up could be spent more constructively.
Public sessions of the Gathering will commence at 10:15 a.m. Monday, May 19, in the Hilton Washington Dulles Airport, 13869 Park Center Road, Herndon, VA 20171.
Additional information, and links for registering for the entire event or for individual sessions, can be found here.
Categories: News • In this article: ecumenism, National Council of Churches of Christ (USA), USA Transmis : 10 mars 2014	• Lien permanente : ecu.net/?p=7504
Catégorie : News • Dans cet article : ecumenism, National Council of Churches of Christ (USA), USA
URL: http://ecumenism.net/2014/03/ncc-first-christian-unity-gathering.htm


text:
CAMPUSPEAK announces members of new board of advisors
Categorized: News
CAMPUSPEAK announced today that seven members have been selected to serve on the company's first Board of Advisors. The new members include two higher education professionals, a business professional, an association professional, two CAMPUSPEAK speakers, and a CAMPUSPEAK facilitator. The members will begin their service on June 23, 2015. Their terms will last through Dec 31, 2016.
The new board members are: Dr. Kevin Bailey, Kristen Hadeed, Karen Katz, Monica Lee Miranda, Marc Mores, Margarita Perez, and James Robilotta.
"Our advisors' range of experience and skills are a perfect addition to our team," said CAMPUSPEAK president David Stollman. "It is truly exciting to have such great people committed to helping CAMPUSPEAK. I trust that their expertise and leadership will present new opportunities for our continued growth."
Dr. Kevin Bailey has served as the Vice President of Student Affairs at the University of West Florida since 2010. He has had an extensive range of experience for over 20 years serving roles at Tulane University, Millersville University, Bowling Green State University, UNC Charlotte, and Indiana University of Pennsylvania. Bailey has been a leader in the student affairs field through his active involvement in numerous Higher Ed associations including Southern Association of College Student Affairs, ACPA, and NASPA. Bailey has remained an active member of his fraternity Kappa Alpha Psi at the local, regional, and national level.
Kristen Hadeed graduated from the University of Florida in 2010 with a degree in Finance. When she was a junior in college, Kristen founded Student Maid™, a cleaning business that only employs students who maintain a GPA of 3.5 or higher. The company now has two locations in Florida and more than 500 employees during peak seasons. In addition to working with her team to grow the Student Maid™ brand nationwide, Kristen launched a tech company that is developing scheduling apps to revolutionize the service industry. Kristen currently lives in Florida where she frequently guest lectures at the University of Florida. She is known for her TEDx talk "How to Retire by 20" and recently landed a book deal in hopes to continue spreading her message.
Karen Katz is the Chief Executive Officer of Phi Delta Epsilon Medical Fraternity and currently serves as the Professional Fraternity Association President. She has an undergraduate degree in Theatre Management from the University of Evansville and a graduate degree in College Student Personnel from Western Illinois University. Since 2000 she has been advising fraternities and sororities. Karen volunteers for Chi Omega on the National Recruitment Team and as the University of Miami Recruitment Advisor. She was the Fraternity and Sorority Life Advisor at Florida International University (FIU) for four years where she starting volunteering for numerous fraternities including Sigma Phi Epsilon. She has been a lead facilitator for the North-American Interfraternity Conference's Undergraduate Interfraternity Institute since 2004.
Monica Lee Miranda received her bachelor's in Spanish with a minor in Psychology and her master's in Educational Administration and Policy Studies from the University at Albany. She is now in the dissertation stage of her doctorate degree in Educational Leadership at University of Rochester. Monica currently serves as the Director of the Center for Student Involvement at the University of South Florida and is the past President of the Association of Fraternity/Sorority Advisors and is a member of Omega Phi Beta sorority.
Marc Mores graduated from Iowa State University in 1995 and received a M.Ed. from Xavier University in 2004. He currently serves as the Executive Vice President of James R. Favor & Company running daily operations of the Denver office and serving as underwriter for their Lloyd's of London fraternity insurance program. Additionally, he serves on the Board of Directors of HazingPrevention.Org and remains active with his fraternity, Phi Delta Theta, in local and national volunteer capacities. Having worked for the fraternity in roles ranging from regional director to Associate Executive Vice President for 13 years, he is frequently called on by his, and many other fraternal organizations, to provide educational programs at leadership conventions.
Margarita Perez serves as the Dean of Students at the University of Louisiana, Lafayette. A native of the Crescent City, Margarita fits right in with the Ragin' Cajuns having received her M.Ed. from the University of New Orleans and undergraduate degree from Loyola University. Margarita served 12 years at Spring Hill College in the roles of Associate Dean of Student, Assistant Vice President, and then Interim Vice President of Student Affairs. Prior to Spring Hill College, Margarita held positions in Student Life at Newcomb College at Tulane and Residence Life at Southeastern Louisiana University.
James Robilotta grew up in Sayville, NY and attended the University of North Carolina, Wilmington for undergrad. After getting a B.S. in Marine Biology, he earned his Masters of Counselor Education: Student Affairs Practice in Higher Education from Clemson University. While there he also became a proud brother of Iota Phi Theta Fraternity, Inc. James worked as a student affairs professional for over 11 years in residential life and campus activities. James is now a professional speaker and a personal coach. James has been keynoting corporate and collegiate conferences and facilitating staff retreats since 2002, and has won "Top Program" honors on state, regional, and national levels.
Since 1999, CAMPUSPEAK has provided transformative learning experiences through its keynote speakers, interactive workshops, consulting, and facilitation services. Partnering with campuses and higher education organizations across the country, CAMPUSPEAK offers programming to educate and inspire students for success in their college years and beyond. Learn more at campuspeak.com.


text:
Q: How to show Master Detail information in a new dialog box c# wpf I have a datagrid with a list of "JobItem" objects, and when I select an item from this list, I want to be able to open it up in a new window and be able to edit its properties and save it.  I have the databinding part of it, I just don't understand how I can pass the object instance that I selected from datagrid, and have its information populated in the new window.  I found the exact project in codeproject that did what I wanted to do and looked at the code but I still could not figure out who this object instance was passed.  The article is:
http://www.codeproject.com/Articles/332615/WPF-Master-Details-MVVM-Application

A: Cast the ListViewItem to the object.
`(ObjectType)YourListView.SelectedItem`

will give you an instance of the class, if that's what you're asking.
You can then pass it to the edit form through it's constructor. If you pass it by value, you'll have to write some update code in the original form to "merge" the changes.
I think if you pass it by reference, it'll update the listview on it's on if you have it set up correctly with INotifyPropertyChanged. You may want to test that to make sure, though. I'm not 100% on that.

redpajama_set_name: RedPajamaStackExchange


text:
New Mobile App Helps Companies Overcome Language Barriers When Going Global
globalization strategy, Localization, messaging app, translation, translator
Language: it is the hidden side of globalization, the less glamorous but often crucial piece of a company's globalization strategy.
As companies like Huawei and Amazon gone global, they have relied more and more on localization companies to internationalize their products and services. In turn, localization companies have had to innovate the tools they use to accommodate this new demand.
Stepes, a chat-based messaging app, aims to give the localization and translation industry with a much-needed boost while making translation more accessible to both translators and businesses.
"Localization is so much more than translation," explains Carl Yao, an executive vice president at CSOFT International, a localization company headquartered in Beijing and San Francisco. "That's why we created Stepes, a mobile translation app that is the first to use a texting-based interface to help with localization."
Stepes, mobile app works like this. First, a large translation project gets divided up into smaller components and given to individual translators. Companies can view the profiles and selected translation samples of all Stepes translators, who come from a worldwide pool of 100,000+ translators assembled by Stepes' partner site, TermWiki.com.
Stepes – and the emergence of various online human translation services – represents a shift in strategy among businesses. Spurred on by globalization and slower domestic consumer spending, Asian companies and firms have sought to expand their global market shares. Going global however, requires translating all their products, interfaces, and websites into tens of different languages simultaneously. In particular, many companies looking to globalize are in the technology and e-commerce, raising new challenges like creating appealing product names, translating technical manuals, and constantly updating online content in several languages at once. As a result, the translation and localization industry is predicted to grow to $39 billion by 2018, and a large portion of that growth will come from Asian companies looking to expand globally.
"There's incredible potential for innovation in localization right now," said CSOFT's Yao. "That's why we need to be able to scale-up translation to meet globalization needs."
Currently, there are only 2100,000 professional translators worldwide for nearly 6,500 languages. The vast mismatch in the supply and demand for translation has resulted in much higher translation costs for businesses, slower turnaround time for projects, and a lack of coverage for lesser known languages. Traditional translation tools are usually desktop-based and require some technical knowledge of software, slowing down the translation process and also turning away many would-be translators.
Yet Yao notes that over half the world speaks at least two languages, if not more; that's up to 3.65 billion people who have the potential to be translators, at least part time. Stepes targets these bilingual speakers and hopes that by providing free and easy to use translation tools directly through smartphones, more people will begin translating in their spare time. Collectively, even if each person only contributed a few sentences at a time, Yao believes that our translation capacity would still grow exponentially to meet true translation needs today.
Yao calls this expansion of our translation capacity a "big translation" approach. By harnessing the collective language talents of people around the world, he hopes to drive down translation costs for businesses while also empowering bilingual people to translate in their free time and earn money. In this way, Stepes users can "share" and pool language abilities that would otherwise have gone underutilized to tackle large translation projects for globalizing businesses.
Apps like AirBnB and Uber have already (successfully) tried this sharing economy model with travel accommodations and taxi rides. Could it work for translation? That is still unclear, but Stepes may be our best bet so far.
Read the original article on AsiaToday
Stepes, world's first chat-based translation app, now available on iTunes and Google Play
Stepes Applies Mobile Messaging Interface to Translation
Why Ecommerce Companies Like Amazon Need Stepes Big Translation Solutions
Miley Thomas
Mobile Translation for a Mobile Society
Lizzy Broek
Why global trade wars will accelerate the arrival of globalization 2.0, creating tremendous opportunities for the translation industry
Why GDPR Is a Boon for Online Translation Services like Stepes
China's Decision to Lift Restrictions on Car Manufacturing by Foreign Automakers Promises Tremendous Opportunities for Language Translation Services
Tom Armstrong
A new life in the Philippines
Marie Adams
Why crypto currencies like Bitcoin and Ripple are ideal for the global translation market
Vicarious Trauma a Lesson in Empathy
Emily Hargreaves


200 hours yoga teacher training course is one of our most popular at our yoga ashram in Goa and Dharamsala. It is a Yoga Alliance recognized certification that makes you a certified yoga teacher on completion.
Weekly and bi-weekly meditation retreats are the second offering of our Goa and Dharamsala based yoga school. These yoga retreats are perfect for people who want to experience yoga without joining a professional course.
Both 200 hours yoga teacher training course and yoga retreats at our yoga school at highly affordable and focused on bringing out the best within you. Rest assured, Bird of Paradise Training Yoga will give you the best package for your yoga goals!
Ready to dive into a yogic and spiritual experience? Email us at emmasmobilebeautyservices@hotmail.co.uk / Pradeepkentura@gmail.com or call at +919205853278 to discuss your TTC and retreats queries with our talented team.
What will you learn at TTC?


Inspired by the smash-hit multi-million selling albums Dreamboats and Petticoats One, Two, Three, Four and Five, the West End sell out sensation Dreamboats and Petticoats The Musical features some of the greatest hit songs of the Rock 'n' Roll era. The dazzling success of the first five albums in the Dreamboats and Petticoats series sent the message loud and clear, after several weeks at the Number One spot in the compilation charts, the Great British public were saying that they didn't just want to listen to pure nostalgia: they'd love to see it as well.
In 1961 emotions run high as young musicians Norman and Bobby compete to win a national song writing competition - and, more importantly, the attention of the gorgeous Sue! But when Bobby discovers that shy Laura is no slouch on the piano, love and rock 'n' roll fame beckons.
Featuring classic tracks from Roy Orbison, The Shadows, Eddie Cochran, Billy Fury, and many more including Let's Dance, To Know Him Is To Love Him, Shaking All Over, Bobby's Girl, Little Town Flirt, Only Sixteen, Runaround Sue, Happy Birthday Sweet 16, Let It Be Me, Great Pretender, C'Mon Everybody, Let's Twist Again and many more hits from music's golden era!