c‐Fos expression in dopaminergic and GABAergic neurons of the ventral mesencephalic tegmentum after paradoxical sleep deprivation and recovery
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Abstract Evidence suggests that dopaminergic neurons of the ventral mesencephalic tegmentum (VMT) could be important for paradoxical sleep (PS). Here, we examined whether dopamine (DA) and adjacent γ‐aminobutyric acid (GABA)‐synthesizing neurons are active in association with PS recovery as compared to PS deprivation or control conditions in different groups of rats by using c‐Fos expression as a reflection of neural activity, combined with dual immunostaining for tyrosine hydroxylase (TH) or glutamic acid decarboxylase (GAD). Numbers of TH + /c‐Fos + neurons in the substantia nigra (SN) were not significantly different across groups, whereas those in the ventral tegmental area (VTA) were significantly different and greatest in PS recovery. Numbers of GAD + /c‐Fos + neurons in both VTA and SN were greatest in PS recovery. Thus, DA neuronal activity does not appear to be suppressed by local GABAergic neuronal activity during PS but might be altered in pattern by this inhibitory as well as other excitatory, particularly cholinergic, inputs such as to allow DA VTA neurons to become maximally active during PS and thereby contribute to the unique physiological and cognitive aspects of that state.Keywords:
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Abstract The ventral tegmental area (VTA), the origin of dopaminergic cell bodies that comprise the mesocorticolimibic DA system, is widely implicated in drug and natural reward, cognition, and several psychiatric disorders. In addition to dopaminergic neurons, this region is populated by GABAergic neurons, which both regulate the firing of their dopaminergic counterparts and send projections throughout the brain. Although the dopaminergic neurons of the VTA have been extensively characterized neuroanatomically, much less is known about the GABAergic neurons in this region. Recent data suggest that the rostro‐caudal topographic organization of these GABAergic neurons may correspond to their ability to regulate drug reward. In the present study, we used immunohistochemical techniques to examine the frequency and topography of GABAergic neurons throughout the rostro‐caudal axis of the VTA and the extent to which they coexpress other proteins, including tyrosine hydroxylase (a marker of DA neurons), cholecystokinin, parvalbumin, calretinin, and calbindin d 28k. Synapse 61:87–95, 2007. © 2006 Wiley‐Liss, Inc.
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Objective To explore the changes of tyrosine hydroxylase neurons in mesencephalic ventral tegmental area (VTA)of Parkinson disease (PD)rats. Methods Stereotaxic injection of 6-OHDA into the left substantia nigra pars compacta (SNC) and ventral tegmental area (VTA) of rats to develop PD rat model, The behavioral changes induced by apomorphine were observed at 4,7,14,21,28 days after the injection of 6-OHDA; neuronal alteration of left mesencephalic VTA were studied adopting Nissl's staining, and the changes of tyrosine hydroxylase neurons were analyzed by tyrosine hydroxylase (TH) immunohistochemistry and image analysis. Results The PD rats showed abnormal rotary behavior, in the rotation, the start time of PD rats shortened gradually, the continuous time lengthened, and the rotary rate speeded by degrees. The rotary behavior became invariable until 2 weeks after 6-OHDA injection; Serious neuronal alteration in left (the injured side)VTA of PD rats were observed by Nissl's staining; The number of tyrosine hydroxylase neurons in left (the injured side)VTA of PD rats, decreased significantly compared to right side throughout the test time. As to the number of tyrosine hydroxylase neurons in left (the injured side)VTA, 2 or 4 weeks after 6-OHDA injection decreased significantly compared to 1 week after 6-OHDA injection in PD group (P0.05). Conclusion The PD rats showed serious neuronal alteration in the injured VTA, tyrosine hydroxylase neurons were lost significantly in the injured VTA of PD rats, Mesencephalic VTA tyrosine hydroxylase neurons take part in the pathologic changes of PD rats.
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The VTA is a critical component of the brain reward circuitry and necessary for processing reward as well as drug reward. DARPP‐32, first discovered in the striatum represents an exciting target for drug‐induced neuroplasticity in the brain, yet the cellular localization of DARPP‐32 in the VTA has not been confirmed. We explored the possibility whether VTA GABAergic neurons, the main local inhibitor of VTA dopamine neurons, express DARPP‐32. To confirm whether a subset of these DARPP‐32‐immunoreactive terminals arise from local GABAergic interneurons, we used a double immunoflurescence confocal microscopy technique using antibodies against mu opioid receptors (as the marker for GABAergic neurons) and DARPP‐32 to visualize GABAergic neurons expressing DARPP‐32. Our preliminary experiments confirmed that some subset of mu opioid receptor‐expressing GABA neurons also expresses DARPP‐32, suggesting that the source of DARPP‐32 in the VTA may arise from some subset of local GABA neurons in the VTA. To our knowledge, this is the first demonstration of the cellular localization of DARPP‐32 in the VTA. Supported by R075OU‐DOD/USUHS and PhRMA Research Starter Grants.
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AIM To determine the effects of tetrahydroprotoberberines (THPB) on functions of neurons in the ventral tegmental area (VTA) of morphine dependent rats. METHODS Rats received daily increasing dose of morphine for 10 d (5, 10, 15, 20, 25, 30, 35, 40, 45, 50 mg·kg -1 , respectively, ip, twice a day) to establish morphine dependence, then were given normal saline (NS) or THPB 30 mg·kg -1 , ip, twice daily for 12 d or 30 d. Control group received ip NS all the time. The levels of glial fibrillary acidic protein(GFAP) and tyrosine hydroxylase(TH) were determined as immunohistochemical intensity by ABC method and image analysis. RESULTS Levels of GFAP and TH in the VTA region of morphine dependent rats were higher than those of control group (P0.05),while groups treated with THPB were similar to those of control group (P0.05). CONCLUSION Higher levels of GFAP and TH indicate that morphine dependence induces impairment of dopaminergic functions in the VTA region. THPB treatment can reverse the damage, which suggest that THPB be benefit for clinical use to prevent heroin relapse.
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BACKGROUND: Taking into account the importance of GABAergic brain system research and also the opportunity to achieve specific and accurate results in laboratory studies using immunohistochemical approaches, it seems important to have a reliable method of visualization GABA-synthesizing cells, their projections and synapses, for the morphofunctional analysis of GABAergic system both in normal conditions and in the experimental pathology. AIM: The aim of the study was to visualize analyze GABAergic neurons and synapses within rats brain using three different antibody types against glutamate decarboxylase and to identify the optimal conditions for reaction performing. MATERIALS AND METHODS: The study was performed on paraffin brain tissue sections of 5 adult Wistar rats. Immunohistochemical reactions using three antibody types against glutamate decarboxylase isoform 67 (GAD67) and glutamate decarboxylase isoform 65 (GAD65) were performed. Additional controls on C57/Bl6 mice and Chinchilla rabbits brain samples were also carried out. RESULTS: Antibodies used in the research made it possible to achieve high quality of GABAergic structures visualizing without increasing background staining. At the same time different antibody types are distinct in their efficacy to perform immunohistochemistry reaction on laboratory animal brain tissue samples. By performing additional controls, we discovered that there is necessary to adsorb secondary reagents immunoglobulins in order to eliminate nonspecific staining. It was found that GAD67 and GAD65 distribution in rat forebrain structures is different. It was stated that GAD67 immunohistochemistry most completely reveals GABAergic brain structures compared to GAD65 immunhistochemistry. The possibility of determining morphological features of GABAergic neurons and synaptic terminals, as well as performing quantitative analysis, was demonstrated. CONCLUSIONS: The approach proposed makes it possible to specifically visualize GABAergic structures of the central nervous system of different laboratory animals. This could be useful both in fundamental studies and in pathology research.
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