Amanda Charlesworth, PhD CharlesworthAmanda@uams.edu

I am interested in the molecular changes that drive developmental processes. In the CTN I am investigating the molecular mechanisms that control the developmental decrease in REM sleep. Newborn mammals spend a large proportion of time in REM sleep and by adulthood, this has substantially decreased. The reticular activating system is thought to regulate REM sleep but it is not known how. We have been investigating the hypothesis that neurons in the reticular activating system are electrically coupled via gap junctions. We have found that the neuronal gap junction protein, connexin-36, is expressed in the SubCoeruleus nucleus of the reticular activating system (see figure below). Additionally, connexin-36 levels decrease during development, a decrease that mirrors the decrease in REM sleep. The mechanisms behind the decrease in expression of connexin-36 are being investigated.
 

Connexin 36 expression and protein levels over age.  A. Ratio of Cx 36 mRNA vs each of three housekeeping genes, Enolase (dark gray), Hprt (light gray) and Gapdh (black), which did not differ from each other in developmental expression.  Mean and SE of 3 mesopontine tegmenti of 7 day, 17 day and adult rats.  Note the gradual decrease in Cx 36 expression in development but with significant levels present in adults.  B. Cx 36 protein levels in representative mesopontine tegmenti of 7 day, 17 day and adult rats.  Levels decreased gradually from 7 to 17 days to adult, such that longer exposures (Connexin-36 hi-exp) were required to visualize protein levels in adults.  Blot was reprobed for actin protein levels to verify equal loading.  C. Photomicrograph of 400 um sagittal slice through a 30 day medial brainstem showing punched region (1 mm diameter) ventral to the Locus Coeruleus (the punch was centered anterior to the 7th nerve but included the nerve as it descended) in the SubC region.  D. Cx 36 protein levels were normalized to Actin in slices containing the SubC from rats from 4 litters aged 10 days and 30 days.  Note the decrease in protein levels across the developmental decrease in REM sleep.  E.  Cx 36 protein from these pooled samples from 10 day and 30 rats normalized to Actin loading.  Actin protein levels are shown to verify equal loading.