Title:
[Selective stimulations and lesions of the rat brain nuclei as the models for research of the human sleep pathology mechanisms].

Abstract:
Many complex behavioral phenomena such as sleep can not be explained without multidisciplinary experimental approach, and complementay approaches in the animal models "in vivo" and human studies. Electrophysiological, pharmacological, anatomical and immunohistochemical techniques, and particularly stereotaxically guided local nanovolume microinjection technique, enable us to selectively stimulate and lesion the brain nuclei or their specific neuronal subpopulation, and to reslove the mechanisms of certain brain structure regulatory role, and its afferent-efferent connectivity within the brain. Local stereotaxically guided nanovolume microinjection technique enable us to investigate in animals the brain nulcei functional topography with a resolution of < or = 10 microM, and at a level of 300 microM of effective radius within the brain tissue "in vivo". The advantage of local glutamate or DL- homocysteic acid microinjection stimulation or local excitotoxic (glutamate, ibotenic acid, IgG saporin) microinjection lesion over electrical stimulation/lesion of the same neuronal population are that they reduces the likelihood of activation/lesion of fibers of passage. Much of our knowledge of the sleep neuronal substrates is based on animal studies primarly in cat and rat. Selective pharmacological stimulation of the pedunculopontine tegmentum (PPT) in freely moving rat, using glutamate microinjection, proved that excitation of its cholinergic part is necessary for induction of wakefulness or REM (Datta S, 2001). Local nanovolume glutamate microinjection into PPT of anesthetized rats (Saponjić et al, 2003a) additionally evidenced P-wave and respiratory regulating neuronal subpopulation within the cholinergic compartment of PPT (apneogenic neuronal zone). Local microinjection of serotonin and noradrenaline into cholinergic PPT apneogenic zone evidenced their opposed impact through PPT on breathing, in contrast to their convergent regulatory role in behavioral state control (Saponjić et al., 2005a). Also, selective pharmacological stimulation by microinjection of DL-homocysteic acid defined four neuronal micro-circuitry approximately 500 microm in lenght of breathing-related neurons within the ventral respiratory group of medulla oblongata, which when stimulated produce different effects on respiratory rate, rhythm and amplitude, and on blood pressure. This study was the first high resolution study in order to understand anatomical and functional neuronal system organization (Monnier et al., 2003). Recently, local glutamate microinjection stimulation technique enabled detailed functional topography of respiratory, cardiovascular and pontine-wave responses within the PPT (Topchiy et al., 2010). Discovery of "flip-flop" switch for REM sleep control is based on the experiments in rats using local stereotaxically guided microinjection of excitotoxins (ibotenic acid, IgG saporin), and the anterograde and retrograde tracers for selective lesion, and identifying "REM-off" and "REM- on" regions and their afferent-efferent connections, and for identifying pathways for REM atonia and REM EEG activation (Lu et al., 2006). Recently, selective lesion of SLD part of "REM-on" region in rat established an animal model of RBD, as well as a selective ibotenic acid lesion of PC part of "REM-on" region abolished theta during REM (Lu et al., 200; Anaclet et al., 2010). Selective ablation targeted to pre-Bötzinger complex neurons of ventrolateral respiratory group of medulla in rat induced REM related respiratory disorder up to 10 days, when this respiratory disorder became spreaded to all sleep phases, and even during wakefulness, due to long-lasting intermitent hypoxia, and an increase of the threshold for hypoxia/hypercapnea induced arousal response (McKay et al., 2005). Human development, maturation, healthy aging and many neurological diseases are associated with profound changes in sleep/wake states distribution and with variety of the sleep-related behavioral disorders. Sleep and sleep-related respiratory disorders (insomnia, hypersomnia, parasomnias, excessive nocturnal motor activity, circadian sleep-wake rhythm disturbances, respiratory dysrhythmias, RBD) are very frequently unnoticed in patients with neurodegenerative diseases (Boeve et al., 2007; Whitwell et al., 2007). Alzheimer's and Parkinson's disease (AD, PD) are the most common neurodegenerative diseases, with prevalence of 0.5-1%; increasing to 1-3% for Parkinson, and up to 50% for Alzheimer's disease in ages over 69 (Nussbaum and Christopher, 2003). In spite of a long knowledge of their clinical description and brain pathology (lesions of the NB cholinergic neurons in basal forebrain, dopaminergic neurons in substantia nigra, etc.), they remain incurable with only limited success in temporal amelioration of their symptoms. Clinical symptoms first appear at 65-69 years on average, but there are indications that subclinical features may start many years earlier. Patients with REM-sleep behavior disorder (RBD) face close to a 20% 5-year risk of developing PD or dementia, and that risk rises to more than 40% after 10 years, and exceeds 50% after 12 years. Human studies evidenced that sleep/wake cycle disturbance, as no cognitive symptom of dementia, precedes on average 3 years before the clinical diagnosis of the AD (Simic et al., 2009), and that RBD, precedes as symptom the onset of motor and cognitive disturbances by years or decades. AD and PD involve the selective loss of specific neuronal populations within the brain. RBD in those patients reflects an underlying synucleinopathy, with presence of the alpha-synuclein protein pathology within the REM sleep-related regulatory structures of the dorsal midbrain and pons at the onset of disease, with ascending pattern of neurodegeneration progression from brainstem to basal areas of the brain (Whitwell et al., 2007; Simic et al., 2009: Raggi and Ferri, 2010). On the base of hypothesis that basal forebrain cholinergic system plays an important role in the etiology of the most common neurodegenerative diseases of elderly (AD, PD), the lesion of the nucleus basalis in rat presents the most utilized "in vivo" animal model to study the disorders of cortical cholinergic innervation, and its impact on higher central nervous system functions. Our knowledge of the neural substrates for sleep/wake states and sleep-related behavior disorders regulation in health and the diseases, over more than 50 years of sleep research, is based on animal models, pharmacotherapy, central nervous system lesions, and the neuropathological studies in humans. Today we have many complementary animal models of human sleep pathology, and further work in fundamental multidisciplinary and clinical research between sleep and neurodegenerative disease investigators is promising to enable us understand normal and abnormal sleep, and may provide new insights into preventive or disease-altering approaches for therapy. Obviously counseling and prevention of AD or PD would be highly enriched by the development of a practical, sensitive and reliable methodology of detecting those patients with RBD, or other sleep disorders, who are at risk for developing AD or PD.