Sleep deprivation and sleep recovery modifies connexin36 and connexin43 protein levels in rat brain Javier Franco-Pe ´ rez a , Paola Ballesteros-Zebadu ´a b , Edith A. Ferna ´ ndez-Figueroa c , Isabel Ruiz-Olmedo d , Pablo Reyes-Grajeda d and Carlos Paz a Gap junctional communication is mainly mediated by connexin36 and connexin43 in neurons and astrocytes, respectively. It has been suggested that connexin36 allows electrical coupling between neurons whereas connexin43 participates in several process including release of ATP. It was recently reported that blockage of gap junctional communication mediated by connexin36 can disrupt the sleep architecture of the rat. However, there is no experimental approach about effects of sleep deprivation on connexins expression. Therefore, we examined in adult male Wistar rats whether protein levels of connexin36 and connexin43 change in pons, hypothalamus, and frontal cortex after 24 h of total sleep deprivation and 4 h of sleep recovery. Western blot revealed that total sleep deprivation significantly decreases the levels of connexin36 in the hypothalamus and this decrease maintains after sleep recovery. Meanwhile, connexin43 is not altered by total sleep deprivation but interestingly the sleep recovery period induces an increase of this connexin. These results suggest that electrical coupling between hypothalamic neurons could be altered by sleep deprivation and that sleep recovery drives changes in connexin43 expression probably as a mechanism related to ATP release and energy regulation during sleep. NeuroReport 23:103–107  c 2012 Wolters Kluwer Health | Lippincott Williams & Wilkins. NeuroReport 2012, 23:103–107 Keywords: connexin36, connexin43, sleep deprivation a Department of Neurophysiology, b Medical Physics Laboratory, National Institute of Neurology and Neurosurgery, c Department of Experimental Medicine, National Autonomous University of Mexico and d Medical Proteomics Unit, National Institute of Genomic Medicine, Me ´xico D.F., Me ´ xico Correspondence to Javier Franco-Pe ´ rez, PhD, Department of Neurophysiology, National Institute of Neurology and Neurosurgery M.V.S., Insurgentes Sur No. 3877, Col. La Fama, C.P. 14269 Me ´xico D.F., Me ´ xico Tel: + 52 555 606 3822; fax: + 52 555 424 0808; e-mail: jfranco@innn.edu.mx Received 20 October 2011 accepted 25 October 2011 Introduction Gap junctions (GJ) are clusters of channels composed of protein subunits named connexins (Cx’s) that allow direct communication between cells. These transmem- branal channels contribute to denominated gap junctional communication because they allow fast passage of ions and biological molecules as well as electrical coupling or synchronous firing of coupled neurons [1]. First studies about Cx’s expression demonstrated that Cx36 is highly expressed in neurons [2], even in-situ hybridization analysis has shown significant levels of Cx36 mRNA present in neurons of several regions in both developing and adult brain [3]. In contrast, some studies have reported that astrocytes can functionally couple to other astrocytes by means of GJ [4]. Moreover, based on ultrastructural and immunostaining experiments it is now well recognized that Cx43 is expressed mainly in astrocytes [5] at the earliest developmental stages and its expression remains high through adulthood [6]. There are reports regarding the incidence of GJ in brain regions that have been implicated in the regulation of sleep–wake cycle. Specifically, electrophysiological and molecular evidence has been reported about the presence of electrical coupling and both Cx36 gene expression and protein in the mesopontine tegmentum, specifically in the pedunculopontine and SubCoeruleus nuclei [7,8]. In addition, there are studies that recognize the presence of Cx36 and Cx43 in GJ of the locus coeruleus (LC) of adult rodents [9,10]. Given that pedunculopontine, SubCoer- uleus and LC are some of the brain regions involved in the basic circuitry underlying the regulation of sleep and wakefulness [11], it has been proposed that electrical coupling based on the presence of GJ in these structures could be a novel mechanism that contributes for the regulation of the sleep–wake cycle. According to this hypothesis, a study from our laboratory showed that blockage of GJ induces a significant increase of wakefulness at the expense of sleep [12]. However, this is a topic that has not been fully investigated and at present it has not been determined whether disruption of normal patterns of sleep could alter the expression of the structural substrate of GJ. Because total sleep deprivation (TSD) and sleep recovery (SR) are useful tools to analyze effects of sleep loss and subsequent sleep rebound, the aim of this study was to determine the changes of Cx36 and Cx43 protein levels in some rat brain regions after TSD and SR. Methods Eighteen male Wistar rats (280–320 g) were used for this study. Animals were maintained under controlled condi- tions (8:00–20:00 h light : 20:00–8:00 h darkness) and with Sleep 103 0959-4965  c 2012 Wolters Kluwer Health | Lippincott Williams & Wilkins DOI: 10.1097/WNR.0b013e32834e8fcb Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.