We are very proud to announce that MacICT has been successful in our application for a 2014 'Computer Science for High School' grant from Google!
This annual program promotes computer science education by helping to equip educators with the skills and resources they need to teach computer science and computational thinking concepts in fun and relevant ways. Globally, this program has already trained more than 12,000 teachers and reached over 600,000 students. Read all about it here.
Earlier this year, MacICT applied for one of these grants based on the need we identified for workshops aimed at teachers inspired by the Maker movement. This year they opened it up for us to target Primary as well as Secondary teachers, and we will be running a workshop for teachers from Kindergarten to Year 12 called 'MakerDay C.R.E.A.T.E! – How to Create Reality with Electronics, Art and Technology in Education!'.
Despite ongoing rhetoric around the need for "Twenty First Century" educators, a computer science skills shortage has been identified by the employment sector. This skills shortage is evident at both an international and local level and has been attributed to the slide in the number of computer skills training courses offered within schools over the last two decades. In turn, this has manifested into a shortfall of skilled workers in the computer science industry. The current number of blue and white-collar CS workers is now disproportionate to the demand of an increasingly digital world.
There is a high need to engage students in technological skills and for students to learn how to design and create new digital technologies and applications yet to be invented. With this in mind, half of the proposed "MakerDay C.R.E.A.T.E! How to Create Reality with Electronics, Art and Technology in Education" (MakerDay C.R.E.A.T.E.) workshop is to be spent introducing to select digital programming languages suitable for the age group of students they teach.
In parallel to the deficit of skilled CS worker lies a shortage of skills in manufacturing and trade, particularly in the industries of woodwork, metalwork, electronics and engineering. Due partly to a similar decline in courses offered within schools over the same period, many students of today have had limited exposure in learning how to creatively design and make things. The remaining half of the proposed workshop's focus therefore is to expose teachers and their students to the exciting and internally gratifying process of learning how to create objects that have both form and function, through understanding basic engineering principles, electronic circuits and robotics.
Teachers will see that an opportunity exists for greater student engagement across several Key Learning Areas of the school curriculum, while also offering students realistic vocational choices in computer science, engineering or robotics before they leave school, go to University or begin employment.
In addition, the one day "MakerDay C.R.E.A.T.E! How to Create Reality with Electronics, Art and Technology in Education" professional learning workshop will provide K – 10 teachers with a range of exciting pedagogical strategies, ideas and resources. These will help introduce the concept of Maker Education as a way for teachers to design learning opportunities that develop confidence, creativity and spark an interest in science, technology, engineering, maths and the arts. This workshop will show teachers how students can create technology through inventing solutions to real world problems.
Due to a growing awareness of the computing requirements of the draft Acara Digital Technologies curriculum and the demands of the NSW BOS syllabuses for the Australian curriculum, teachers are looking for ways to integrate ICT in meaningful and authentic ways to meet cross curricula outcomes. Increasingly, teachers in Australian schools are recognising the need to integrate computing into learning but lack the knowledge and confidence to do so in innovative and creative ways.
A deeper understanding of learning through design, as a pedagogical approach, situated in an inquiry-based context where students are cast as problem solvers involved in co-constructing knowledge, ideating, inventing, prototyping and iterating.
A range of teaching strategies, project ideas, resources that integrate computing with electronics and mechanics to create engaging, authentic learning experiences for children that spark creativity and develop confidence.
The knowledge, skills and support necessary to introduce programming, electronics and engineering and robotics to meet the design and computing requirements of the draft 2013 Acara Technologies Curriculum, as well as the demands of the NSW Board Of Studies, Teaching and Educational Standards (BOSTES) for the National Curriculum. There will be a particular emphasis on developing computational, design, creative, critical and systems thinking.
Click here to read more and enrol now!


The Asians held a 9-4 margin in aces. They also made fewer errors 21 to 29 by the opponents. Miyuki Horie was the best scorer for the winning team with 18 points, followed by Air Kurogo with 16. Beatriz Carvlho and Nyeme Alexandre had 18 points each one in the loss side.
Brazil dominated the start of the game and lead the technical times out by 8-7 and 16-12. Japan's defence could not contain the explosive attacks of Brazil´s Nyeme Alexandre, who closed the first set 25-21.
In the second set was Japan that took the lead of the game, with a technical and aggressive serve controlled the actions of The South American and won the set 25-21 with an outstanding performance of the wing spiker Ai Kurogo.
Due to problems in reception Brazil struggled to structure their attacks. The Asians were always leading the score (8-4, 16-14). With great performances of MiyuNakagawa and Miyu ki Horie closed the third set 25-23.
The Japanese used to play with the support of local fans, feeling confident and quickly took control of the fourth set 17-10. Brazil tried to stay in the game, however the multiple combinations of Japan ended the final set by 25- 18.


The National Agricultural Statistic Service's Nov. 10 crop production report had several people in the Mid-South rice country shaking their heads in disbelief.
The report pegged this year's U.S. rice crop at 203.5 million hundredweight with average yield estimated at 6,959 pounds per acre. The estimate for yield and production were much higher than many anticipated — given problems with the crop in Arkansas, which produces about 45 percent of the U.S. crop.
NASS estimated average yield for Arkansas at 6,850 pounds per acre on 1.345 million acres, but dropped the yield only 50 pounds per acre from its October estimate. By comparison, Mississippi's yields, at 7,000 pounds per acre on 229,000 acres were dropped 200 pounds since October.
If the average yield in Arkansas this year were off just 10 bushels (560 pounds) from last year's yield of 7,130 pounds, average yield for the state would be 6,570 pounds per acre, 280 pounds per acre under the NASS estimate. That would lower Arkansas rice production by 6 million hundredweight, to 88 million hundredweight. The U.S. rice crop would decline from 203 million hundredweight to 197 million hundredweight.
Last month, NASS had to correct its Oct. 10 crop production report because its acreage figures for dry edible beans, canola, corn, sorghum, soybeans, and sunflower did not match those on file with the Farm Service Agency. No mention was made of rice, however. Wilson says the discrepancy for rice acres still exists, but added that NASS is probably being "conservative" in making revisions.
Nonetheless, Wilson says that much of this year's Mid-South rice crop was in trouble from the beginning. "The crop was about three weeks late. Then, the fact that the crop was late meant we had so much of the crop in the field when the hurricanes (Gustav and Ike) came through. If we had been in a normal harvest window, we would have had half the crop out of the field before the hurricanes ever came through. We were set for less yield from the get go because we didn't get the crop planted on time.
"Growers tell me they are down from last year anywhere from 10 bushels to 20 bushels, farm average, which is 10 percent to 15 percent," Wilson said.
All of Smith's rice is stored on-farm, and he plans to hold it until prices improve. "The economic current is running swiftly downhill and everything is caught up in it. I think the true fundamentals of the rice market, if they were known, would be bullish.


We are a group of real estate investors seeking a means to meet and interact with other like minded investors and related professionals, so we can all do more deals and conduct more business.
Events: Networking events are open to the public, with a small $10 entrance fee per person to cover our costs. We encourage a relaxed, informal and fun atmosphere.
Local Speakers: Monthly events will include lots of ways to interact and topics of interest presented by local speakers, with no selling speakers. If a speaker is not local to New England, they will be speaking on a topic of interest to real estate investors and not selling a program or course.
No Selling Speakers: Speakers will not sell books, tapes, CD's, courses, bootcamps or mentoring programs at our networking meetings.
No Seeking "Investors": Attendees are welcome to bring deals to buy and sell, and to introduce themselves to the group. No presentation of "investment opportunities" such as real estate syndications. No seeking "Investors or Partners". This is an SEC violation. No offering of "mentorship programs". No offering hard money.
No Multi-level or Network Marketing: Multi-level marketing programs are not allowed in this venue. Please do not solicit other attendees that you meet for multi-level programs during or after the meeting.
Patronize the Venue: We depend on the welcome of our venue providers, and would encourage you to patronize the restaurant during the event. Join us before and after the event for a snack, meal or beverage.
No Memberships, no Dues: There are no membership dues and therefore no long list of discounts and other perks as benefits of membership.
Black Diamond: Tim Murray, $3 Billion invested in Worcester! Be there!


Greetings again from the darkness. Don't mistake this for either the Franco Nero (1966) or Jamie Foxx (2012) movie. This latest from writer/director Etienne Comar centers on Django Reinhardt, one of the most talented and influential musicians of the twentieth century. Based on the novel "Folles de Django" by Alexis Salatko, the story follows the challenges of his escape from German-occupied France.
He is already a renowned (and enigmatic) performer when the film kicks off in 1943 Ardennes, as Django and his band are being contracted by the Nazis to tour and entertain the troops. Of course, he refuses to sign the contract and tour under their terms with limit the style of music he can play. Because of this, Django and his family must flee and disappear underground, while they plan an escape to Switzerland.
His musical influence proliferated the area, and his influence and respect is clear at each step of his travels. In fact, it's the musical pieces and segments that really stand out here. Reda Kateb (A PROPHET, 2009) gives a terrific and expressive performance as Django, but the musical portions are so outstanding, that we find ourselves not as engaged in the personal saga of escape as we should. Clearly, the war and Nazis are a threat, and when Django says "I'm a musician. It's what I do", that serves as his admission that he takes an apolitical stance and does not envision himself as a hero to the people.
As a driving force behind European jazz, and being such an influence on so many guitar players, Django's legacy is something other than as a war icon. The film certainly could have benefitted from more attention to either how his music gained popularity, or what drove him to avoid any political notoriety until it was too late for many of his fellow Gypsies. Admittedly, his escape was crucial and led to his 1945 score, "Requiem for Gypsy Brothers", of which his conducting leads to the most emotional moment of the film.


text:
August 2022 EdNSW
27 July 2022 | August 2022, Editors NSW, Newsletters
From the committee
By Paul Anderson and Russell Noakes
The EdNSW committee met on 12 July for its monthly meeting via Zoom, with three apologies.
Minutes of the June committee meeting were accepted and actions from previous meetings reviewed.
Written reports received included:
Accreditation Board delegate
branch financial report for May 2022
budget commentary with consolidated outcomes to end May and preliminary estimates for June 2022
IPEd Board update
speaker presentations.
We finalised the agenda and related arrangements for our Annual Branch Meeting (ABM) and social quiz on 2 August 2022, and invite all branch members to attend this free Zoom event. The main business is to declare the result of the branch committee election, to constitute the new committee and to present the branch activity program and budget.
Thanks to those branch members who responded to our calls for committee nominations. IIt is still not too late if you do wish to join the committee. There will be another call to fill vacant committee positions on the night, during the ABM.
There was no speaker presentation in July, with our winter dinner in Sydney that month instead.
Our next speaker presentation is on 6 September. Aditya Vasudevan from Arts Law will present on "Defamation law for editors". Arts Law will host the Zoom presentation and the event will be externally advertised in addition to our normal promotion. We expect a high number of registrations from both members and affiliates. So book early.
Two IPEd Associate members and one Student member joined the branch in June. The branch had 367 active financial members at 30 June 2022.
Our financial position is sound and we're operating within the budget guidelines.
The next committee meeting – the first meeting of the new committee for 2022–23 – is scheduled for 9 August 2022.
Many EdNSW events will continue to be held via Zoom.
Please check your email for the latest information on any event.
Please make bookings via the IPEd Events webpage.
Details of our next events are below.
There are no speaker events planned for August due to the ABM.
Editors New South Wales annual branch meeting
Details: NSW members are invited to join the committee to hear about the branch and activities over the past year, see in the new committee and stay for a social gathering including a quiz. BYO beverage of choice for the social part of the meeting.
Bookings: Details have been sent to members via email.
"Defamation law for editors" with Aditya Vasudevan
Date: Tuesday 6 September 2022
Time: 7.00 pm AEST
Details: This presentation will introduce the Arts Law Centre of Australia, then give an overview of the basics of defamation law, before discussing how editors can manage the risks of defamation litigation. The scenario of an editor editing someone's personal memoir will be used. Participants will have opportunities to ask questions about hypothetical situations. The presentation will be hosted as a webinar by Arts Law.
Presenter: Aditya Vasudevan is a solicitor at the Arts Law Centre of Australia, the country's only community legal centre dedicated to the arts. He regularly advises writers of all types about defamation, copyright, publishing contracts and a range of other issues. Aditya studied Law and Arts at the University of Auckland and co-edited the student magazine there. He started his legal career at Chapman Tripp in New Zealand, working on a number of high-profile defamation cases. Before joining Arts Law, he also worked at Gilbert & Tobin in Australia in the intellectual property team. In his spare time, Aditya writes essays and short stories, and is a dedicated reader of literary fiction.
Government departments and editors
Date: Tuesday 4 October 2022, 7.00 pm AEDT (Note: Daylight saving starts in NSW, ACT, VIC, TAS and SA on Sunday 2 October 2022).
Details: A presentation on the editing needs and practices of government departments
Presenter: David Seale, A/g Assistant Secretary Communication, Australian Department of Education, Skills and Employment.
Bookings will be publicised closer to the event.

redpajama_set_name: RedPajamaCommonCrawl


text:
Volume 222, Issue Suppl_1
Basil el Jundi,
Basil el Jundi
Almut Kelber,
Almut Kelber
Barbara Webb
Behavioural basis of olfactory imprinting
Neuronal basis of olfactory imprinting
REVIEW| 06 February 2019
Behavioural and neuronal basis of olfactory imprinting and kin recognition in larval fish
Gabriele Gerlach
Gabriele Gerlach *
of Biology and Environmental Sciences, Carl von Ossietzky University Oldenburg
Helmholtz Institute for Functional Marine Biodiversity Oldenburg (HIFMB)
Centre of Excellence for Coral Reef Studies and School of Marine and Tropical Biology, James Cook University
*Author for correspondence (gabriele.gerlach@uni-oldenburg.de)
Kristin Tietje,
Kristin Tietje
Daniela Biechl,
Daniela Biechl
Graduate School of Systemic Neurosciences & Department Biology II, Ludwig-Maximilians-Universität Munich
82152 Planegg-Martinsried
Iori Namekawa
Friedrich Miescher Institute for Biomedical Research
Gregor Schalm,
Gregor Schalm
Astrid Sulmann
Deutsche Forschungsgemeinschaft (NO AWARD)
J Exp Biol (2019) 222 (Suppl_1): jeb189746.
A related article has been published: Navigation: from animal behaviour to guiding principles
Gabriele Gerlach, Kristin Tietje, Daniela Biechl, Iori Namekawa, Gregor Schalm, Astrid Sulmann, Basil el Jundi, Almut Kelber, Barbara Webb; Behavioural and neuronal basis of olfactory imprinting and kin recognition in larval fish. J Exp Biol 6 February 2019; 222 (Suppl_1): jeb189746. doi: https://doi.org/10.1242/jeb.189746
Imprinting is a specific form of long-term memory of a cue acquired during a sensitive phase of development. To ensure that organisms memorize the right cue, the learning process must happen during a specific short time period, mostly soon after hatching, which should end before irrelevant or misleading signals are encountered. A well-known case of olfactory imprinting in the aquatic environment is that of the anadromous Atlantic and Pacific salmon, which prefer the olfactory cues of natal rivers to which they return after migrating several years in the open ocean. Recent research has shown that olfactory imprinting and olfactory guided navigation in the marine realm are far more common than previously assumed. Here, we present evidence for the involvement of olfactory imprinting in the navigation behaviour of coral reef fish, which prefer their home reef odour over that of other reefs. Two main olfactory imprinting processes can be differentiated: (1) imprinting on environmental cues and (2) imprinting on chemical compounds released by kin, which is based on genetic relatedness among conspecifics. While the first process allows for plasticity, so that organisms can imprint on a variety of chemical signals, the latter seems to be restricted to specific genetically determined kin signals. We focus on the second, elucidating the behavioural and neuronal basis of the imprinting process on kin cues using larval zebrafish (Danio rerio) as a model. Our data suggest that the process of imprinting is not confined to the central nervous system but also triggers some changes in the olfactory epithelium.
Coral reef fish, Larval dispersal, Orientation, MHC peptides, Olfaction, Crypt cell
Olfactory guided navigation in coral reef fish
Like many aquatic organisms, coral reef fish show a dual life stage, where settled adults produce dispersing larvae. At hatching, planktonic larvae drift away from the reef to spend a species-dependent time (larval dispersal duration) in the open ocean – probably to avoid high predation in the reef. These millimetre-sized larvae quickly (within a few days of dispersal) turn into relatively powerful juvenile swimmers, reaching swimming speeds of several centimetres per second (Fisher et al., 2000; Fisher and Wilson, 2004; Stobutzki and Bellwood, 1994, 1997). Until recently, the distribution and settlement of coral reef fish were assumed to be purely driven by currents and stochastic storm events. However, the persistence of marine populations at small isolated oceanic islands requires that a significant number of juveniles return to the natal habitat after their pelagic dispersal phase (Robertson, 2001). Here, and even in less isolated habitats, self-recruitment and natal homing has been assumed to be (far) greater than purely planktonic dispersal would predict from modelling approaches (Armsworth, 2000; Cowen et al., 2000; Staaterman and Paris, 2014; Wolanski et al., 1997). We used population genetic analysis to demonstrate that up to 60% of juvenile cardinalfish, Ostorhinchus doederleini, could be assigned to the adult reef population where they were about to settle (Gerlach et al., 2007a, 2016). Natal homing of other species, e.g. clownfish juveniles (Amphiprion spp.) was also confirmed by mark–recapture studies, otolith tagging and microchemistry studies (Jones et al., 1999; Swearer et al., 1999; Thorrold et al., 2006).
As it is not possible to track larvae in the ocean, dispersal distances are based on catching larvae in plankton tows and by modelling approaches using the pelagic larval duration as a proxy (but see Weersing and Toonen, 2009). Dispersal distances of coral reef fish larvae are assumed to be shorter than 150 km depending on the species (Burgess et al., 2007; Paris and Cowen, 2004). Despite a potentially wide distribution, numbers of homing coral reef fish juveniles are far higher than expected by random movement. Orientation capabilities could help them to find their way back to their natal reefs.
Finding the way back to a natal reef, river or beach after week- or year-long dispersal might require a learning and memory process involving time-dependent specific parameters of this natal place. For orientation-guided homing to natal reefs, coral reef fish larvae must remember sensory parameters that they experienced directly after hatching, which is the only time in which they can obtain reliable 'home cues', as larvae – with fully developed olfactory and visual sensory systems – start dispersing into the open ocean on the night of hatching.
To explain navigation over long distances, we provided evidence that the juvenile cardinalfish (O.doederleini) can use time-compensated sun compass orientation during the day (Mouritsen et al., 2013) and a magnetic compass at night (Bottesch et al., 2016). For orientation at closer distances, there is evidence that reef fish larvae can also respond to acoustic cues for orientation (Radford et al., 2010, 2011). Additionally, we have documented a pronounced importance of olfaction in the homing of coral reef fish larvae (Atema et al., 2002; Gerlach et al., 2007a; Paris et al., 2013). Coral reef fishes such as O. doederleini, Pomacentrus coelestis and several other species of apogonid and pomacentrid juveniles were shown to prefer the smell of water collected from a reef to that of open ocean water (Atema et al., 2002). Both species were capable of distinguishing between chemical cues from reefs within the Capricorn/Bunker reef group and preferred the smell of water from the reef where they were caught over that of other nearby reefs (Gerlach et al., 2007a). We further demonstrated that the preference for home reef odour did not switch even when larvae were exposed to other reef odours for several days (Miller-Sims et al., 2011). Therefore, we concluded that coral reef fish larvae had imprinted on olfactory cues of their home reefs, similar to the well-known olfactory imprinting of salmon on chemical cues of natal rivers (Hasler et al., 1978; Scholz et al., 1973, 1976). However, the robust experimental manipulations that demonstrate an ability of reef fish to imprint upon artificial odours in a similar way to salmonids have yet to be performed.
Olfactory imprinting and olfactory guided navigation in teleosts
Imprinting is a specific form of long-term memory enabling organisms to recognize specific cues from the environment and conspecifics. It might enable offspring to find their mothers among many other females or help animals to remember and recognize environmental conditions that they experienced shortly after hatching or birth. To ensure that organisms memorize the right cue, the learning process must happen during a specific short time period, which should end before the organism encounters irrelevant signals from unrelated conspecifics or environments. One well-known example of a visual imprinting process is Konrad Lorenz's geese experiment (Lorenz, 1935) in which he showed that goslings imprint on the first moving object that they experience after hatching, which they take to be their mother. Many imprinting processes are involving olfactory cues, probably because in social contexts chemical cues can be taken as reliable 'honest' signals and manipulating one's own body smell is very difficult.
The timing of imprinting has been studied extensively in salmonids, and there is strong evidence that salmon imprint on environmental cues of their natal water during the parr–smolt transformation (PST) when they adapt from living in freshwater to living in seawater (for review, see Dittman and Quinn, 1996). There is more recent evidence that some species of Pacific salmon, e.g. juvenile pink salmon (Oncorhynchusgorbuscha) and others, can imprint prior to the PST at a very young age (Bett et al., 2016; Dittman et al., 2015). Bett et al. (2016) published an excellent review on olfactory imprinting and olfactory guided navigation during spawning migrations in different salmon species and lampreys, elucidating imprinting cues and underlying mechanisms. However, there is also evidence that salmon imprint magnetically on their home area and this most likely brings them close to the home stream, with olfactory cues used mainly in the final part of the spawning migration (Putman et al., 2014, 2013).
In addition to natal water, anadromous fish also respond to water that is conditioned by conspecifics, which they remember and recognize years later, when they search for their natal river (for review, see Keefer and Caudill, 2014). There is evidence that salmonids are even capable of population-level discrimination (Courtenay et al., 1997; Doving et al., 1974; Groot et al., 1986; McBride et al., 1964; Nordeng, 1971; Nordeng and Bratland, 2006; Olsén, 1989; Quinn and Tolson, 1986; Selset and Doving, 1980), and even discrimination of siblings from non-siblings (Quinn and Busack, 1985; Quinn and Hara, 1986; Winberg and Ols, 1992). But such kin-structured associations are not confined to salmonids. Genetic analyses have shown that in numerous vertebrate and invertebrate species, populations do not consist of a random mix of individuals but of groups of related individuals of full- or half-siblings; kin associations have been demonstrated in many teleost species (for review, see Gerlach and Hinz, 2012; Selwyn et al., 2016). To identify even unfamiliar kin, organisms must imprint on specific cues of kin, which they match with those expressed by unknown conspecifics (phenotype matching) (Hepper, 1986; Tang-Martinez, 2001). Therefore, our findings (Gerlach et al., 2008; Gerlach and Lysiak, 2006; Hinz et al., 2013a,b) on the mechanisms underpinning olfactory imprinting in zebrafish (Danio rerio) might apply to a whole variety of species.
Olfactory imprinting on conspecific and kin cues in zebrafish
While selective advantages of kin recognition were studied in numerous species, the behavioural and neurobiological basis of the imprinting process are much less understood. To elucidate the behavioural and neuronal basis of the imprinting process, we took advantage of the model organism zebrafish (D. rerio), which imprint on an olfactory and visual template of kin early in development, leading to a long-term memory expressed as an olfactory-based differentiation between kin and non-kin (Hinz et al., 2013a).
Zebrafish raised in kin groups (siblings) identify even unfamiliar kin later in life, a process called 'phenotype matching' (Hepper, 1986). We found that larval and juvenile zebrafish preferred olfactory cues of kin when they were given the choice between water in which kin (siblings) or non-kin were kept; larvae were tested in a two-channel 'Atema' choice flume (Mann et al., 2003). The olfactory preference for kin changed with sexual maturity: adult females preferred the odour blend of unfamiliar non-related males over unfamiliar brothers, while adult males were attracted to all females, related and unrelated (Gerlach and Lysiak, 2006).
We raised larvae under different conditions to identify the time window of imprinting (see Fig. 1F). As experimentally isolated larvae (kept in glass beakers) did not express any olfactory preference in the Atema choice flume test, we concluded that they must be exposed to siblings during development. The sensitive period for olfactory templates of kin is limited to 24 h at 6 days post-fertilization (dpf) (Gerlach et al., 2008) (Fig. 1A). When larvae were exposed to kin at other days for 24 h, but not at 6 dpf, they did not express any kin preference later in life (Fig. 1F). We also raised single larvae (hatching occurs at 4–5 dpf at a temperature of 25°C) in separate beakers and could imprint them when adding kin water at 6 dpf (Fig. 1B). Without such olfactory cues, they did not express any preference for kin (Fig. 1C,E). Interestingly, the application of non-kin water at 6 dpf did not result in a preference for non-kin odour (Fig. 1G). Therefore, zebrafish larvae must experience kin odour at 6 dpf for imprinting, and they cannot be imprinted on olfactory cues of non-kin.
Summary of behavioural olfactory kin preference tests. A–O are modified from Hinz et al., 2012, 2013a,b. The diagrams on the left illustrate the raising conditions, which are described in the next two columns (vis., visual cues; olf., olfactory cues). 'Test conditions' describe chemical cues that larvae experienced during the test. 'Olfactory preference' indicates whether larvae expressed a statistically significant preference for kin or no preference; see 'Olfactory imprinting on conspecific and kin cues in zebrafish' for details. MHC II, major histocompatibility complex (class II).
Placing the glass beakers containing single larvae directly next to each other was very important, because it turned out that a visual template was essential for the imprinting and later olfactory-based kin recognition process. Without visual exposure to other kin, there was no kin preference and recognition later in life (Fig. 1D). Interestingly, visual cues of same-aged non-kin did not evoke imprinting (Fig. 1G). Again, we identified the time period for visual imprinting, which turned out to happen at 5 dpf (Hinz et al., 2013a). Both olfactory and visual cues were essential and larvae did not express any kin recognition when the cues were given at any other time than 5 dpf (visual cue) and 6 dpf (olfactory cue) or when originating from non-kin. These findings indicate that a genetic predisposition exists that results in imprinting when larvae experience the right cues.
Our results about the necessity of visual kin cues were intriguing. How can a larva know what its siblings look like when it has never seen any larvae before? What makes them know that the other larvae outside its own beaker are relatives? To understand this phenomenon, we raised single fertilized eggs/larvae in glass beakers surrounded by mirrors – we added holding water of kin at 6 dpf and obtained strong kin preference (Fig. 1H). Without adding kin odour, we did not observe kin recognition (Fig. 1I). We concluded that self-matching on chemical cues is not possible, but self-matching on visual cues is possible.
We wondered whether fish did not imprint on their own body odour because the concentration of urine from a single larva is too low to trigger imprinting. To test this possibility, we kept two full siblings in twice as much water as a single larva, to obtain the same concentration, and found that concentration had no effect: imprinting was observed when two siblings were kept together but not when a larva was kept individually, although a view of additional siblings outside the beaker was possible in both conditions.
To understand the visual part of imprinting, we took photos from the lateral side of larvae and compared the pigment pattern within and between sibling groups; this showed that related larvae share a similar pigmentation pattern (Hinz et al., 2012). The highest similarity among related larvae was found in the pigmentation pattern of the eye, indicating that the process is based on genetic disposition rather than direct comparison of own-body pigmentation with that of others. Larvae in a natural environment cannot see their own eye pigmentation in contrast to larvae in the above-mentioned mirror experiments.
Advantage of kin groups
What is the selective force for olfactory imprinting and resulting kin recognition in zebrafish larvae? Larvae raised in groups of full-siblings showed an increased growth rate of 33% compared with larvae raised with non-kin (Gerlach et al., 2007b); however, we could not observe any differences in aggression or foraging. Therefore, association with kin and faster growth rates might lead to enhanced fitness by earlier reproduction. In Atlantic salmon and rainbow trout, kin group members had more foraging opportunities and experienced significantly greater weight gain than non-kin group members (Brown and Brown, 1996). In three-spined sticklebacks (Gasterosteus aculeatus), Frommen and Bakker (2004) suggested that shoaling with kin reduces competition, allows for more stable dominance hierarchies and increases fitness of relatives.
MHC class II peptide ligands as chemical signals evoking olfactory imprinting and kin recognition
In vertebrates, much effort has been made to reveal which cues are used to identify kin. The kin signal must fulfil two attributes: it must be sufficiently unique to avoid mistakes and it has to be stable in order to be recognized over space and time. In the aquatic environment, these are non-volatile olfactory cues. The expression of genes of the major histocompatibility complex (MHC) provides individuals with a unique body odour that is more similar among individuals with increasing genetic relatedness (Apanius et al., 1997; Boehm and Zufall, 2006; Penn, 2002; Singh, 2001). Imprinting on MHC and other relatedness cues released via urine, mucus and gills plays a major role in kin recognition in fish and other aquatic species (for review, see Gerlach and Hinz, 2012).
The olfactory preference of individual zebrafish larvae appears to be defined at least in part by their MHC class II genotype. We used a combination of artificial MHC peptides to study their potential role in imprinting (Hinz et al., 2013b). First, we searched for sibling groups, which developed a preference for a specific set of five randomly chosen synthetic MHC peptides when they had been exposed to these peptides during the critical period for imprinting. We found one zebrafish line (OL1, one pair and their offspring) that showed olfactory preference after exposure to these peptides at 6 dpf (Hinz et al., 2013b) (Fig. 1J). This exposure resulted not only in a subsequent preference for the MHC peptide mix but also in a preference for kin water (Fig. 1K), suggesting that the chosen MHC peptides are identical or similar to components of the kin-specific odour cues that are relevant for imprinting in this line (Fig. 1).
We further examined whether zebrafish can imprint on olfactory cues of other unrelated zebrafish that bear the same MHC genotype. The similarity of MHC class II but not MHC class I alleles between two zebrafish larvae corresponded to imprinting. Larvae did not discriminate between water derived from fish with identical MHC class II alleles even though they differed in genetic relatedness (Hinz et al., 2013b) (Fig. 1L–O). Hence, the MHC class II genotype appears to code for important olfactory cues for kin recognition and imprinting in zebrafish.
Interestingly, MHC class II allele sharing led to similarity not only in olfactory cues but also in pigmentation pattern. The degree of allele sharing of MHC class II genes defined body, rump, tail and iris pigmentation (Hinz et al., 2012).
We concluded that the MHC class II genotype could represent the underlying mechanism carrying the information about kin and non-kin and, thus, be the basis of the genetic predisposition (Hinz et al., 2013b).
Olfactory system and its signal transduction in fish
Unlike that in mammals, the olfactory system of the zebrafish as a teleost consists of only one main olfactory organ (Hansen et al., 2004; Hansen and Zeiske, 1993, 1998). Olfactory chemoreception is dependent on the binding of an odorant to its corresponding receptors, located on microvilli or cilia of olfactory sensory neurons (OSNs), with subsequent signal transmission into the central nervous system. Olfactory receptors (ORs) are located on the dendrites of OSNs embedded in the olfactory epithelium (OE) (see Fig. 2). Two major populations of OSNs are present in fish – microvillous OSNs (mOSNs) and ciliated OSNs (cOSNs) – resembling OSNs present in tetrapod vomeronasal and main olfactory systems (Eisthen, 1997; Korsching, 2016). Zebrafish feature two more OSN types: crypt cells (Ahuja et al., 2013; Hansen and Finger, 2000; Kress et al., 2015) and kappe cells (Ahuja et al., 2014) – both are named after their peculiar morphology and are so far only known to be present in fish (see Fig. 2). Odorant information is transduced into electrical signals inside the OSNs. OSNs in the nose express one olfactory receptor out of a large repertoire and project directly to the first olfactory processing centre in the brain, the olfactory bulb (OB), where many of them converge into fewer glomeruli (primary olfactory pathway) (Yoshihara, 2009). Bulbar projections further transmit odorant information into different areas of the telencephalon and diencephalon (secondary olfactory pathway) (Yoshihara, 2009). For example, in carp, mOSNs are mainly tuned towards food-related odours, whereas cOSNs play a role in mediating the alarm reaction, and crypt cells are suggested to be involved in reproduction (Hamdani et al., 2001, 2008; Hamdani and Døving, 2002, 2006). Electrophysiological data from Hansen et al. (2003) in channel catfish suggest that all OSN types respond to amino acids, with cOSNs additionally responding to bile salts, whereas mOSNs also respond to nucleotides. In goldfish, mOSNs are assumed to respond preferentially to amino acids (Speca et al., 1999). In transgenic zebrafish, blocking of synaptic transmission in distinct populations of mOSNs abolished attractive behavioural responses to amino acids (Koide et al., 2009), which is in line with the findings of other studies (Lipschitz and Michel, 2002). Amino acids and nucleotides are typically indicative of food whereas bile salts are considered as social odorants as bile salt profiles within a teleost family and order show high similarity (Hagey et al., 2010). However, a general conclusion on teleost OSN tuning is hard to make as the response profiles of OSNs differ, as do bulbar projection patterns across species (Bazáes et al., 2013).
Zebrafish odorant receptors and olfactory sensory neurons (OSNs). (A) Olfactory epithelial positions of zebrafish OSNs. Combinatorial analysis of calcium-binding protein (CBP) expression reveals at least eight subpopulations of OSNs. Bold font indicates the largest of three subpopulations of ciliated OSNs (parvalbumin/calbindin/calretinin, PV/CB/CR; red). Four subpopulations of microvillous OSNs (mOSNs, dark blue) express PV. CBP S100 is specifically present in all crypt cells (green) and a minor subpopulation of mOSNs, which also express PV (shown again on the right of crypt cells in light blue). Kappe neurons (purple) probably express PV and CB and probably correspond to the PV/CB-mOSN population. (B) Zebrafish olfactory sensory cell types defined by morphology and CBPs. Schematic drawing of a cross-section through a larval zebrafish olfactory epithelium and olfactory bulb of one body side. The mediodorsal glomerular field (mdG) is indicated by six glomeruli (1–6). A hypothetical representation of the crypt cells (green) projecting to mdG2 is shown. Additional axonal input to this glomerulus comes from S100/PV-expressing microvillous OSNs (light blue). A second glomerulus (out of the 6) within the mediodorsal domain, mdG5 (5), receives input from PV/CB-expressing OSNs, probably representing kappe neurons. Also shown are the ventroposterior glomerulus (vpG) and the ventrolateral (VLF) and ventromedial (VLM) glomerular fields subserved by mOSN and cOSN input, respectively. See text and Kress et al. (2015) for more details. (C) Comparison of zebrafish olfactory receptors and OSNs with those in mouse and human (for sources of data, see 'Olfactory system and its signal transduction in fish').
Experience of olfactory cues can induce changes in the olfactory neuronal system
The olfactory system of teleosts enables the recognition and discrimination of a vast number of environmental and social odours. In the aquatic realm, olfactory-based behavioural context includes migration (Yamamoto et al., 2010), association with kin (Hinz et al., 2013b; Olsen et al., 2002), feeding (Miklavc and Valentincic, 2012), predator avoidance (Idler et al., 1956), response to alarm signals (Schreckstoff) (Speedie and Gerlai, 2008) and social interactions such as mating and reproduction (Veyrac et al., 2011). The olfactory system of fish detects and discriminates between structurally different classes of odorants: mainly charged molecules including amino acids, bile acids, nucleotides, steroids and prostaglandins, but also uncharged volatile molecules (Korsching, 2016). Responses to amino acids, bile acids and nucleotides (Carr, 1988) emerge early in development, shortly before hatching (Li et al., 2005). In zebrafish and other teleosts, defined prostaglandins and steroids serve as pheromonal signals prior to mating (Kobayashi et al., 2002; Moore and Waring, 1996; Sorensen et al., 1988; Stacey et al., 1989; Yabuki et al., 2016), and substances released from the skin evoke an alarm response (Jesuthasan and Mathuru, 2008). Most of the behavioural responses to these stimuli arise late in development, but the emergence of physiological and neurobiological responses to these stimuli has not been studied in detail. However, Braubach et al. (2013) provided exciting evidence of an activity-dependent developmental mechanism in the zebrafish olfactory system, which might be important to understand olfactory imprinting. They suggested that imprinting could trigger neuronal changes in the OB and perhaps in the expression of specific olfactory receptor genes dependent on contact with olfactory cues at a specific time during development. Ochs et al. (2017) described the morphology of glomerular patterning in wild Chinook salmon during a temporal window previously shown to be significant for early olfactory imprinting.
Sockeye salmon (Oncorhynchus nerka) that successfully imprinted on l-arginin, as evidenced by adult behaviour, demonstrated increased expression (relative to arginine-naive fish) of the putative arginine receptor mRNA in the OE during key life stages (Dittman et al., 2010). In brains of hatchery-reared juvenile chum salmon (Oncorhynchus keta), thyrotropin-releasing hormone gene expression increased immediately after release from a hatchery into the natal stream, and the expression of the essential NR1 subunit of the N-methyl-d-aspartate receptor (NMDAR) increased during downstream migration (Ueda et al., 2016). The NMDAR is a glutamate receptor channel subtype and mediates most of the fast-excitatory synaptic transmission in the central nervous system. It plays important roles in memory formation and retrieval in fish (Gómez et al., 2006; Kinoshita et al., 2004). Gene expression of salmon gonadotropin-releasing hormone (sGnRH) and NR1 increased in the adult chum salmon brain during homing from the Bering Sea to the natal hatchery (Ueda et al., 2016). However, in contrast to salmonids, in zebrafish, we could not evoke any olfactory preference for a variety of amino acids after larvae were exposed to different concentrations of these amino acids during development (K.T. and G.G., unpublished data). However, zebrafish as a non-migrating fish might imprint on other cues.
Tuning of teleost OSNs and processing of kin odour in the zebrafish OE
To investigate which type of OSN detects kin odour, we mapped neuronal activity following olfactory stimulation visualized by pERK upregulation in OSNs in the OE of 9 dpf zebrafish larvae (Biechl et al., 2016). pERK is widely used to mark neuronal activity in mammals; for example, in the olfactory system following odour stimulation in mice (Mirich et al., 2004). Upon phosphorylation, pERK is translocated into the nucleus of the activated cell to modulate the expression of transcription factors, which in turn regulate the expression of genes involved in neuronal and synaptic plasticity underlying learning and memory. pERK is preferable to other markers, such as immediate early genes (e.g. c-fos or egr1), because of its more rapid activation and its cellular distribution (soma and cell protrusions) (Gao and Ji, 2009; Randlett et al., 2015).
Olfactory stimulation with different odours (food, conspecific odour or E3-medium) showed a differential pattern of activated OSNs within the larval OE (Biechl et al., 2016). High numbers of cOSNs – and a smaller number of mOSNs – were activated in response to food odour exposure, whereas mOSNs almost exclusively showed pERK upregulation following exposure to a conspecific odour (Biechl et al., 2016). Interestingly, crypt cells showed neuronal activation in response to neither food nor conspecific (non-kin) odour. These response patterns were generally consistent with data from other studies, despite great interspecific variability regarding the tuning of OSNs within teleosts (Bazáes et al., 2013).
By rearing larvae isolated in small glass beakers in a larger tank, we either prevented olfactory imprinting on day 6 or allowed the larvae to imprint on their kin by adding kin odour containing water to the beakers (Biechl et al., 2016). The rearing conditions for all larvae were otherwise identical (e.g. changing water, feeding and isolation), precluding other influences on the results. Importantly, despite raising larvae in glass beakers, visual imprinting at 5 dpf was possible because isolated larvae could see their siblings swimming outside their own beakers in the larger tank (see also Hinz et al., 2013a). Additionally, in the subsequent histological assay for pERK we used the calcium-binding protein (CBP) S100 to specifically identify crypt cells. In the crucian carp (Carassius carassius), crypt cells project into the ventral OB from where projection neurons terminate in the lateral part of the medial olfactory tract, which is known to mediate reproductive behaviours (Weltzien et al., 2003). Furthermore, in sexually mature carp, crypt cells were shown to vary in their density as well as location within the OE depending on the season (Hamdani and Døving, 2006; Hamdani et al., 2008). Carp exhibited only a few such cells during winter whereas in the summer spawning season, carp crypt cells were clearly detectable and positioned at the surface of the OE, suggesting that they were involved in carp reproductive behaviour (Hamdani et al., 2008). Sandulescu et al. (2011) demonstrated an interesting time course of zebrafish crypt cell quantity during OE development. Upon first appearance in the zebrafish OE at 4 dpf, the number of crypt cells increased steadily until reaching a peak at 7 dpf. Comparing this growth of crypt cells with the sensitive phase of olfactory imprinting (day 6), it appears as though crypt cell numbers might adapt to the upcoming imprinting event.
Based on these studies, we exposed imprinted and non-imprinted zebrafish larvae at 9 dpf to kin odour; only crypt cells of imprinted larvae showed activation in response to kin odour (Fig. 3C; Biechl et al., 2016). Importantly, this was not due to an absence of this cell type, as crypt cell numbers did not differ between non-imprinted and imprinted larvae (Biechl et al., 2016) (Fig. 3A). Furthermore, microarray data provided no evidence for a down-regulation of ORA4 receptor expression in crypt cells in non-imprinted compared with imprinted zebrafish larvae (K.T. and G.G., unpublished data). Thus, the critical cue, which is obviously contained in kin odour, apparently changed the responsiveness of crypt cells in an unknown manner during the imprinting process. Crypt cells express only a single V1R-like receptor, encoded by the ora4 gene (Oka et al., 2012). The ligand that binds to ORA4 is still unknown; however, based on our data, the ligand of ORA4 is contained in kin odour but it is unlikely that ORA4 is the specific receptor for MHC peptides in zebrafish (Behrens et al., 2014; Boschat et al., 2002; Isogai et al., 2011).
Differential activation of crypt cells with kin odour – effects of olfactory imprinting. (A) Total cell quantity of S100+ mOSNs and crypt cells. Imprinting has no effect on total cell number. (B) Number of S100+/pERK+ mOSNs shown as a percentage of all S100+ mOSNs per larva. Significantly more S100+ mOSNs are activated in imprinted larvae versus non-imprinted control larvae exposed to kin odour. (C) A significantly higher number of crypt cells is activated after kin odour stimulation in imprinted compared with non-imprinted larvae and compared with imprinted control larvae stimulation. No difference in activation was found within non-imprinted larvae. (D) The total number of pERK+, but S100−, ciliated OSNs (cOSNs), mOSNs and crypt cells. Cell activation was similar for all treatments in cOSNs. A significantly higher number of mOSNs was found in imprinted larvae stimulated with kin compared with control stimulation. No S100− crypt cells were observed. Statistical significance: **P<0.01, ***P<0.001. Modified from Biechl et al. (2016).
In addition to crypt cells, a small subpopulation of (S100-negative) mOSNs was shown to respond to kin odour (Fig. 3D). Considering that MHC peptides are a component of kin odour, these mOSNs might express a V2R receptor and bind to such peptides. Regardless, kin odour is composed of a mixture of numerous odorants that signal – besides genetic relatedness – additional social information such as sex and physiological status, and possibly kin recognition is mediated by more than one chemical compound. Thus, a receptor code, generated by the interaction of multiple receptors and activated by more than one ligand, might signal information about relatedness in larval zebrafish. In sum, our studies support the hypothesis that crypt cells as well as a small subpopulation of mOSNs are involved in detecting a kin odour-related signal.
Potential neuronal changes due to olfactory imprinting
Nevitt et al. (1994) hypothesized a model for olfactory imprinting in salmon. They suggested that during sensitive imprinting periods, thyroid hormones stimulate the proliferation of non-specific olfactory sensory neurons. Subsequently, the receptors that are most responsive to the odorants survive and the non-responsive receptors die. This model was based on previous studies that demonstrated odorant-specific sensitization of the peripheral olfactory system in coho salmon (Oncorhynchuskisutch) (Dittman et al., 1997). Their results suggested that exposure of coho salmon to an odorant for 10 days during smolting could result in significant changes in the peripheral sensitivity to that odorant in the spawning adult coho salmon (Dittman et al., 2010, 1997).
We tested whether the expression of odorant receptors and other genes in zebrafish larvae changed during the sensitive phase of olfactory imprinting through a genome-wide transcriptional analysis using microarray chip that included gene-level probe sets for 129 OR, 82 TAAR, 46 OlfC and 6 ORA odorant receptor genes, covering all known ora genes, but missing probe sets for some odorant receptor genes from the TAAR, OR and OlfC group (K.T. and G.G., unpublished results). Microarray analysis showed no differences in gene expression for 263 odorant receptor genes in imprinted versus non-imprinted larvae (K.T. and G.G., unpublished results); however, we might have missed specific odorant receptors especially some TAARs, which are suggested to be associated with the detection of social cues in mice (Ziegler et al., 2002).
Zebrafish larvae exposed to the artificial odorant β-phenethyl alcohol (βPEA) showed an increase in expression of the transcription factor Otx2 inside the OE (Calfún et al., 2016; Harden et al., 2006). The authors suggested that βPEA triggers olfactory imprinting in juvenile zebrafish and further showed that three OR genes (OR111-1, OR115-5, OR125-1) are affected by Otx2 expression.
Our qPCR analysis in 5, 6 and 7 dpf zebrafish larvae (before, during and after olfactory imprinting) revealed no differences in Otx2 expression; the same finding was obtained for imprinted and non-imprinted larvae (K.T., G.S and A.S., unpublished results). In situ hybridization experiments confirmed our findings: no differences in Otx2 expression pattern were observed in 5, 6 and 7 dpf imprinted and non-imprinted larvae.
Spatial distribution of odour responses in the OB
To explore how olfactory cues relevant for imprinting are represented in the zebrafish brain, we measured odour-evoked activity in the OB and adjacent areas by multiphoton calcium imaging (Hinz et al., 2013b). Patterns of input activity across the array of glomeruli are processed by neuronal circuits in the OB that are composed of principal neurons, the mitral cells and multiple types of interneurons. General odorants such as amino acids, bile acids and nucleotides stimulate multiple glomeruli, and stimuli of the same class often evoke responses in overlapping, yet stimulus-specific, subpopulations of OB neurons (Friedrich, 2013).
The spatial distribution of odour responses is usually broad but not entirely random because responses to odours of the same class are biased towards broad sub-regions. For example, responses to amino acids and bile acids are found predominantly in the ventrolateral and the dorsomedial OB, respectively (Doving et al., 1980; Friedrich and Korsching, 1997, 1998a; Yaksi et al., 2009). This coarse functional topography of odour responses is established at early stages of development and maintained throughout adulthood (Li et al., 2005). Pheromones, in contrast, have been reported to activate few glomeruli with high specificity (Friedrich and Korsching, 1998b; Yabuki et al., 2016). Moreover, pheromones and general odorants activate different brain areas downstream of the OB (Yabuki et al., 2016), consistent with the hypothesis that pheromonal signals are detected by specific receptors and control distinct behavioural or endocrine responses.
Neuronal representation of olfactory kin cues in the OB
In mammals, MHC peptides convey information about individuality and stimulate sensory neurons of the main and accessory olfactory systems (Leinders-Zufall et al., 2004; Spehr et al., 2006). Individual sensory neurons respond to multiple peptides, indicating that stimulus identity is encoded by a combination of activity patterns across multiple odorant receptors and glomerular channels. However, the spatial distribution of responses to MHC peptides in the OB has not been analysed in detail. To address this, we mapped odour responses to a mixture of the five MHC peptides that evoked olfactory imprinting and kin recognition in a specific kin group (Hinz et al., 2013b) in the OB by multiphoton calcium imaging in transgenic fish expressing the genetically encoded calcium indicator GCaMP2 under the control of the pan-neuronal elavl3 promoter (Ahrens et al., 2012) (Fig. 4). Because zebrafish larvae are small and transparent, activity could be imaged in multiple focal planes, separated by 10 µm, that covered the entire OB and parts of the telencephalon and habenula in vivo. Results from multiple fish were combined by registration of image data to a common reference brain (Fig. 4D). We found that responses to MHC peptides were distributed and not confined to a specific location, although responses appeared to be particularly dense in a sub-volume of the ventrolateral, amino acid-responsive part of the OB (Fig. 4) (Hinz et al., 2013b).
Spatial distribution of responses to MHC peptides and fish odour. (A,B) Expression of GCaMP2 (greyscale) and odour-evoked relative change in fluorescence (colour; thresholded) in four optical sections of transgenic zebrafish. A depth of z=0 corresponds to the dorsal pole of the olfactory bulb; negative values indicate more ventral locations. Right: maximum projection of responses in 10 optical planes spaced at 10 µm. Stimuli were water conditioned by conspecifics (fish odour; A) and a mix of five MHC peptides (B). All fish were raised in the presence of siblings. In trial- and time-averaged response maps, pixels were thresholded at 2.5 s.d. and contiguous areas of less than 20 pixels (∼2.8×2.8 µm2) were deleted. (C) Intersection of activity patterns evoked by fish odour and the MHC peptide mix. In each optical section, response intensities were summed when relative changes in fluorescence exceeded 2.5 s.d. in response to both stimuli. Right: maximum projection of responses in 10 optical sections spaced at 10 µm. (D) Alignment of images from multiple individuals. The four landmarks L1–L4 were aligned by translation, scaling and rotation of image data. OB, olfactory bulb; Tel, telencephalon; Hb, habenula; A, anterior; P, posterior; R, right; L, left. (E) Depth histogram of optical planes imaged in six fish (10 µm spacing). (F–H) Thresholded and averaged responses to fish odour (F) or to MHC peptide mix (G); (H) intersection. Maximum projections through 10–11 optical planes in each of six fish were aligned and combined by taking maximum values. Optical sections covered the entire olfactory bulb but only parts of the telencephalon and habenula.
Unlike responses to amino acid odours, responses to MHC peptides did not substantially broaden with increasing stimulus concentration (Hinz et al., 2013b), consistent with observations in mammalian sensory neurons (Leinders-Zufall et al., 2004). The best response was observed at very low concentrations of MHC peptide mix (1.25×10−12 mol l−1) (Hinz et al., 2013b). Activity patterns evoked by MHC peptides showed significant overlap with activity evoked by the odour of conspecifics (Fig. 4), but less overlap with activity evoked by food extract (Hinz et al., 2013b), even though food extract is a strong and complex odour. These observations support the hypothesis that MHC peptides are released into the water and detected by the olfactory system of zebrafish. Moreover, unlike pheromones, MHC peptides do not appear to be detected by a small number of narrowly tuned sensory channels. Rather, MHC peptides appear to be represented in a combinatorial fashion by activity patterns across neuronal populations, consistent with observations in the peripheral olfactory system of mammals (Leinders-Zufall et al., 2004; Spehr et al., 2006).
As the observed higher neuronal activity in specific OSNs (crypt cells and mOSNs) in imprinted larvae is not explained by an increased number of these OSNs or a change in odorant receptor gene expression, olfactory imprinting might be explained by a change in sensitivity of the odorant receptor itself. A ligand–MHC protein–odorant receptor interaction might lead to a stronger neuronal activation compared with that in larvae that are exposed to peptide ligands of non-kin odour. Odorant-binding proteins can enhance the solubility of hydrophobic odours and facilitate the transport of odours to receptor sites (Pelosi, 1994). Even if this does not represent a modulation of receptor affinity, it still enhances the possibility of a ligand–receptor interaction and thereby might represent a station of neuronal activity modulation in general.
Our approach to study processes that underlie olfactory imprinting and kin recognition in a non-migratory but much more tractable fish should shed light on the mechanistic underpinnings of imprinting in parental populations in migratory fishes. But olfactory imprinting on kin versus environmental cues seems to differ, as shown by differences in olfactory receptor gene expression.
Some parts of the results and discussion are taken from the PhD thesis of Daniela Biechl (Ludwig Maximilian University, Munich, 2018). We thank Sigrun Korsching for fruitful discussions. We also thank B. Grothe for additional support. We thank Pascal Fieth for bioinformatics support for the microarray gene expression analysis. We are grateful to R. Friedrich and Mario Wullimann for their contributions to the discussion.
Funding was provided by the Deutsche Forschungsgemeinschaft DFG (SPP 1392 Integrative Analysis of Olfaction; GE 842/5-1, 5-2, FR 1667/2-1, 2-2 and WU211/2-1, 2-2).
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Caudill
Homing and straying by anadromous salmonids: a review of mechanisms and rates
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Long-term potentiation in the optic tectum of rainbow trout
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Fish Physiol. Biochem.
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Olfactory neural circuitry for attraction to amino acids revealed by transposon-mediated gene trap approach in zebrafish
Aquatic olfaction
Chemosensory Transduction
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Combinatorial analysis of calcium-binding proteins in larval and adult zebrafish primary olfactory system identifies differential olfactory bulb glomerular projection fields
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Breer
MHC class I peptides as chemosensory signals in the vomeronasal organ
Yaksi
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Early development of functional spatial maps in the zebrafish olfactory bulb
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Amino acid odorants stimulate microvillar sensory neurons
Der Kumpan in der Umwelt des Vogels
J. Ornithol.
Turnell
Kin recognition in juvenile zebrafish (Danio rerio) based on olfactory cues
Biol. Bull.
Olfactory perception in juvenile salmon: II. Conditioned respose of juvenile sockeye salmon (Oncorhynchus nerka) to lake waters
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Chemotopy of amino acids on the olfactory bulb predicts olfactory discrimination capabilities of zebrafish Danio rerio
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How stable are the reef odor preferences of settling reef fish larvae?
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Experience-dependent activation of extracellular signal-related kinase (ERK) in the olfactory bulb
Electrophysiological and endocrinological evidence that F- series prostaglandins function as priming pheromones in mature male Atlantic salmon (Salmo salar) Parr
Sun compass orientation helps coral reef fish larvae return to their natal reef
W. J.
Evidence for a peripheral olfactory memory in imprinted salmon
Is local orientation of anadromous fishes determined by pheromones?
Homing experiments with parr, smolt and residents of anadromous Arctic char Salvelinus alpinus and brown trout Salmo trutta: transplantation between neighbouring river systems
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Behaviour of mature anadromous char (Salmo alpinus L.) towards odorants produced by smolts of their own population
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Can. J. Fish. Aquat. Sci.
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Population genetics, larval dispersal, and connectivity in marine systems
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Does the lateral bundle of the medial olfactory tract mediate reproductive behavior in male crucian carp?
Winberg
The influence of rearing conditions on the sibling odor preference of juvenile arctic charr, Salvelinus alpinus L
Wolanski
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Yabuki
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Olfactory receptor for prostaglandin F2alpha mediates male fish courtship behavior
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Transformation of odor representations in target areas of the olfactory bulb
Olfactory imprinting of amino acids in lacustrine sockeye salmon
Molecular genetic dissection of the zebrafish olfactory system
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Celebrating 100 years of discovery
We are proud to be celebrating 100 years of discovery in Journal of Experimental Biology. Visit our centenary webpage to find out more about how we are marking this historic milestone.
Craig Franklin launches our centenary celebrations
Editor-in-Chief Craig Franklin reflects on 100 years of JEB and looks forward to our centenary celebrations, including a supplementary special issue, a new early-career researcher interview series and the launch of our latest funding initiatives.
Looking back on the first issue of JEB
Journal of Experimental Biology launched in 1923 as The British Journal of Experimental Biology. As we celebrate our centenary, we look back at that first issue and the zoologists publishing their work in the new journal.
Biology Communication Workshop: Engaging the world in the excitement of research
We are delighted to be sponsoring a Biology Communication Workshop for early-career researchers as part of JEB's centenary celebrations. The workshop focuses on how to effectively communicate your science to other researchers and the public and takes place the day before the CSZ annual meeting, on 14 May 2023. Find out more and apply here.
Mexican fruit flies wave for distraction
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26 Thoughts: Two USMNT prospects are keying the Timbers, and MLS is Back semifinals predictions
August 5, 2020, 3:12 PM ·12 min read
The play of Eryk Williamson (left) and Jeremy Ebobisse has helped propel the Portland Timbers into the semifinals of the MLS is Back Tournament. (Andrew Bershaw/Getty Images)
Doug McIntyre's MLS column, 26 Thoughts, parses through the latest insights and inside info from around American soccer.
Since this column resumed last month alongside the 2020 Major League Soccer season, we've featured, coincidentally, three of the four teams that comprise the semifinal field for the MLS is Back Tournament. The Philadelphia Union. Minnesota United. Orlando City. Now it's time to give the Portland Timbers some long overdue love ahead of their semi against now-favorite Philly on Wednesday night [8 p.m. ET, FS1/TUDN].
The Timbers have been quietly excellent during their entire stay in Central Florida. They went undefeated in a group that included Supporters' Shield holder LAFC, and after getting a little bit lucky in winning on penalties over surprising FC Cincinnati in the round of 16, they beat the wheels off a New York City FC team that Union coach Jim Curtin likes to call best team in the Eastern Conference.
Yet now it's Portland, not the Pigeons, that stands between Philadelphia and next week's grand finale. And a big reason for the Timbers' success over the last few weeks has been the play of two 23-year-old United States national team prospects: striker Jeremy Ebobisse and central midfielder Eryk Williamson.
26 Thoughts
1. Before we get to Ebobisse and Williamson, let's be clear. The Timbers are stacked. Sebastian Blanco has been sensational; for me, he's the tournament MVP so far. The now-34-year-old Diego Valeri, the face of the franchise since he arrived from Lanus is his native Argentina in 2013, came off the bench and scored the winner against NYCFC. Another South American, Peruvian World Cup veteran Andy Polo, sealed the victory with a golazo.
Then there's midfield destroyer Diego Chara, who just played his 300th match with the Timbers and who Curtin called "one of the most under-appreciated great players that our league's ever had" on Monday. Portland also features Chara's younger brother, winger Yimmi Chara. The list goes on. Still, the improvement of the two young Americans has been obvious.
2. "These young players that you mentioned, Jeremy and Eryk, it's been part of the process that we have had with some of the young players in making sure that they develop, that they grow, that they understand that nothing is easy and they have to earn it and work for it," Timbers coach Giovanni Savarese said when I asked about Ebobisse and Williamson during Monday's pre-match media Zoom call. "We're seeing the fruits of that right now, in Erik especially."
3. Savarese wasn't taking anything away from Ebobisse, who scored three goals in just over 200 minutes during the group stage. The France-born, Maryland-raised striker enjoyed what many considered a breakout 2019, a year that began with his first USMNT cap and ended with him playing in all 34 MLS regular season matches for the Timbers and scoring 11 goals despite being deployed mostly on the wings.
4. Over the last month, though, Williamson's game has taken an even bigger leap forward. The former University of Maryland playmaker's future in MLS seemed up in the air when his rights were traded from D.C. United to Portland in early 2018.
"We believe Eryk is capable of further growth and development, and has the potential to become a quality player at the next level," Savarese said at the time. But Williamson hadn't played a minute for the Timbers first team before being loaned to Portuguese side CD Santa Clara that August. Playing time was nonexistent overseas, too.
5. "It didn't go as expected but I learned a lot," Williamson told Yahoo Sports. "I didn't speak the language, didn't know anyone on the team, didn't know anything about the country. I learned that it's not about how comfortable you are. You have to work hard even if you're not the happiest off the field."
6. Williamson was back in Portland last season but started just three games. He made a brief cameo off the bench in one of the Timbers' two March matches before the coronavirus pandemic shut down the 2020 campaign for more than three months. All of this makes what he's done in Orlando more impressive. The smooth, technical Williamson has already played more minutes over the last four weeks than in his first two seasons as a professional.
7. "He has taken this opportunity to show why he is as indispensable as anyone else on the field," Ebobisse said of Williamson. The two have known each other since they were 13. They grew up playing with and against each other in greater Washington, D.C. and were teammates on the Americans' 2017 U-20 World Cup team that won its group before being knocked out in the quarterfinals. "As a friend I'm really happy for him."
9. And how about this for a comparison? "His ability to dribble through lines, it almost reminds us a little bit of Darlington Nagbe in what he can do for the team," Ebobisse said. High praise indeed. Nagbe, of course, helped the Timbers win MLS Cup in 2015 before earning a second championship with Atlanta United two years ago.
Portland's players compare Eryk Williamson's game to that of former Timbers and current Columbus Crew maestro Darlington Nagbe. (Jeremy Reper/Getty).
10. Williamson is quick to credit Ebobisse for his help. Having a sounding board who had already gone through the inevitable ups and downs all young players face was invaluable. Over the course of our 10-minute conversation, Williamson mentioned more than once how he "looks up to" Ebobisse, who is just four months his senior. "To get to pick his brain, and his opinions on how I'm doing have been massive for my development," Williamson said.
11. Another overlooked part of his path are the 30-plus games Williamson logged with T2 in the second-tier USL. Ebobisse also spent time in the USL, as have USMNT starters such as Tyler Adams and Aaron Long. "Playing in college or with the under-20 national team, it's not the same as playing in a professional game," Williamson said. "It allowed me to get the repetitions and experience I needed."
12. Said Savarese: "In Eryk, now we see the product of three years of work, especially on the side of commitment, understanding and being more focused in the things he has to do.
"Talent, he always has had," the coach continued. "Now he's applying that talent with great work, and that's why he's been very important in this tournament."
13. Savarese took the Timbers to the final in 2018, his first year at the helm. He made the playoffs last year despite starting the season with the longest road trip in MLS history as Providence Park was expanded. Over the years, coaches and GM-types have often told me that it takes three years to build a championship team in MLS."It might take three years, it might take two years," he said. "It takes time to make sure that you put in your work, you put in the culture that you want."
14. Ebobisse says the former Venezuelan international began to build that culture from the start. "Since the day he got here, Gio established a good understanding and rapport with players," he said. "Just on a human level, I think everybody agrees that he's a great person and that makes you wanna fight not only for your teammates but also for the coaching staff and the club."
15. Ebobisse an impressive guy. In addition to emerging as one of the top American goal-scorers in MLS, he's not afraid to speak out about social issues. He's on the board of the Black Players for Change coalition that was formed in the wake of George Floyd's May 25 death in Minnesota police custody to combat systemic racism.
16. The group's efforts have been front and center during the MLS is Back Tournament. Just about every player and coach has been wearing the Black Lives Matter t-shirts designed by Union midfielder Warren Creavalle, one of more than 70 members of BPFC. (The excellent Tom Marshall has the story behind those shirts, which are now on sale to the public.) I asked Ebobisse how the coalition can sustain momentum after the event ends.
"I love history, so I understand that these moments are followed by periods of backlash against whatever the movement accomplished," he said. "Ultimately where we hope to make an impact is off the field., Whether that's front offices, the league office, or in our local markets and our academies. Those are all going to be out of the public spotlight.
"I'm starting to see what we're going through in this country almost as a third reconstruction after the Civil War and then the civil rights movement. We're thinking — I'm thinking — we have a window of years. We have to go for it. And that starts with being organized and making sure that everything you do is fully thought-out, so that people take you seriously. If you create the mechanisms for change during that time period, that can outlast any backlash that comes."
17. I mean, damn. You don't normally hear athletes speak so eloquently about such serious subject matter, let alone one under 25. For a while, the Duke University-educated Ebobisse's friends and family actually worried about his willingness to be candid.
"Before I was drafted people were saying that maybe I shouldn't speak out, that I might be jeopardizing my career," he said. "I told them I can't stop myself. And now I'm among a group of fellow professional athletes who feel passionately about similar issues. We have been able to combine our voices. Hopefully that will make us louder and we can influence the public into at least understanding the experience that we're trying to share and not being against that movement. That, to me, is so important."
18. Before we go any further, it must be said that MLS players are some of the most thoughtful, engaging, down-to-earth pro athletes you'll find anywhere. It was a pleasure to chat with the likes of Ebobisse, Williamson, Tesho Akindele, Michael Boxall, Tim Melia, Chris Wondolowski and Graham Zusi over the last month. Some I've known for years. Others, including our two central figures this week, I'd never spoken to before. But all of them were downright lovely. For me, it's the best thing about covering MLS.
19. One thing that the four remaining teams possess? They really enjoy being around each other. That might seem obvious. It doesn't mean that the other 22 teams don't get along. However, tournaments staged in a neutral location are particularly grueling. The better teams do, the longer they're away from their own beds, from their loved ones. It's a mental grind. With conflicting emotions at play, the team with the best chemistry is often the last one standing.
20. I've been lucky enough to cover multiple World Cups. It's an amazing privilege for any journalist. Here's a dirty little secret, though: At both Brazil 2014 and last summer's Women's World Cup in France, some American scribes were silently hoping the U.S. would get knocked out as soon as possible — so they could go home, too. Some players are the same.
21. Thursday's matchup between Orlando and Minnesota features the delicious storyline of Loons coach Adrian Heath facing the club he took from the third tier to MLS back in 2015 before being fired after that inaugural season. "I have such respect for him," current Lions coach Oscar Pareja said of Heath. The Loons know how much beating Orlando would mean to their coach. But Pareja doesn't expect that to matter. "I don't think you need any extra motivation to compete in a semifinal in Major League Soccer," he said.
On the other hand, it can't hurt. "Everyone's aware of his history with the club," Loons fullback Chase Gasper said of Heath's Orlando link. "We're going to to everything in our power to make sure our coach gets the win in this game.
Minnesota United coach Adrian Heath will face his old team in Orlando City. (Timothy Nwachukwu/Getty)
22. Heath still owns a home in the area. "I'm really looking forward to the game, because you know what my connection with Orlando City is, and the city itself," Heath said. "So it will always be more than just another game for me, because of the respect that I have for the supporters. We built the club together."
23. Heath has plenty of respect for Pareja, too. "We forget how long he's been in the job," he said of Pareja, who was hired during the offseason. "You can see the belief within the group ... we know this will be our toughest game so far."
24. Going 1-for-4 in my quarterfinal predictions (thanks Philly!) isn't going to stop me from handicapping the semis. I still say Philly gets though, although the more experienced Timbers are the bookies' pick by a slight margin. This one is going to penalties, with the Union advancing on the strength of a superior keeper in Andre Blake.
25. Minnesota is the bettors' fave in Thursday's contest. I'm not going against the house twice. Loons win 2-1.
26. Finally, per multiple reports, following the tournament, the league plans to begin playing regular season games in home markets later this month. MLS deserves enormous credit for the way it's pulled off the event in Orlando following lots of skepticism and a shaky start. Along with the NWSL, they showed that playing in a tightly regulated bubble works.
Because of that experience in Orlando, MLS players could be more likely to follow the protocols designed to keep them safe than those in Major League Baseball. However, given the difficulty MLB teams (and for that matter, USL squads) have had staying COVID-19-free in the absence of a bubble, it sure feels like the league is asking for trouble.
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