Abstract
The ventral tegmental area (VTA) is crucial for brain functions, such as voluntary movement and cognition; however, the role of VTA in sleep-wake regulation when directly activated or inhibited remains unknown. In this study, we investigated the effects of activation or inhibition of VTA neurons on sleep-wake behavior using the pharmacogenetic “designer receptors exclusively activated by designer drugs (DREADD)” approach. Immunohistochemistry staining was performed to confirm the microinjection sites, and combined with electrophysiological experiments, to determine whether the VTA neurons were activated or inhibited. The hM3Dq-expressing VTA neurons were excited confirmed by clozapine-N-oxide (CNO)-driven c-Fos expression and firing in patch-clamp recordings; whereas the hM4Di-expressing VTA neurons inhibited by reduction of firing. Compared with controls, the activation of VTA neurons at 9:00 (inactive period) produced a 120.1% increase in the total wakefulness amount for 5 h, whereas NREM and REM sleep were decreased by 62.5 and 92.2%, respectively. Similarly, when VTA neurons were excited at 21:00 (active period), the total wakefulness amount increased 81.5%, while NREM and REM sleep decreased 64.6 and 93.8%, respectively, for 8 h. No difference of the amount and EEG power density of the NREM sleep was observed following the arousal effects of CNO. The inhibition of VTA neurons during active or inactive periods gave rise to no change in the time spent in the wakefulness, REM, and NREM sleep compared with control. The results indicated that VTA neurons activated pharmacogentically played important roles in promoting wakefulness.
Highlights
The ventral tegmental area (VTA) contains dopamine (DA) containing neurons and is traversed by many blood vessels and nerve fibers
◂Fig. 4 Effects of modulation of VTA neurons by designer receptors exclusively activated by designer drugs (DREADD)’ activity during dark period on vigilant states of mice. a–c Hourly amount of wakefulness, rapid eye movement (REM), and non-rapid eye movement (NREM) sleep after CNO administration in the mice microinjected with hSyn-mCherry-AAV10 (n = 6) (a), hSyn-hM3Dq- AAV10 (n = 6) (b), and hSyn- hM4Di-AAV10 (n = 5) (c) into the VTA
The horizontal open and filled bars on the x-axes indicate the 12 h light and 12 h dark periods, respectively. d–g In mice microinjected with hSyn-hM3Dq-AAV10, examples of polygraphic recording and corresponding hypnogram after treatment with saline (d) and CNO (e); time spent in each stage in 8 h (21:00–5:00) following saline and CNO injection (f); amount and EEG power density of NREM sleep in 2 h (5:00–7:00) following CNO-induced arousal (g). *p < 0.05, **p < 0.01, assessed by two-tailed paired Student’s t test the animals were sacrificed and fixed 2 h later for staining
Summary
The ventral tegmental area (VTA) contains dopamine (DA) containing neurons and is traversed by many blood vessels and nerve fibers. Close to its poorly defined borders are found DA and 5-HT containing neurons [1]. The VTA comprises several subdivisions and neurons synthesizing neurotransmitters, such as DA, GABA, or glutamate [2]. The VTA is crucial for brain functions, such as voluntary movement and goaldirected behavior, as well as cognition, emotion, reward, working memory, and decision making [3,4,5,6]. Several lines of evidence suggest that DA signals are closely associated with regulation of sleep-wake activity [8, 9]. Malfunction of the brain DA system is associated with neuropsychopathy, with disruption of sleep as one symptom [10].
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