wake states. Yet MCH does
not appear to exert direct postsynaptic effects on target neurons, including the noradrenergic LC neurons. Previous studies using in situ hybridization showed that MCH neurons synthesize glutamic acid decarboxylase (GAD) and could thus utilize GABA as a neurotransmitter. To determine whether MCH varicosities can release GABA, we examined by fluorescent microscopy in the LC, whether their terminals also contain the vesicular transporter for GABA (VGAT). In dual-immunostained sections, we found that approximately 6% of MCH varicosities was immunopositive for VGAT and a similar proportion for synaptophysin, the presynaptic marker for small synaptic vesicles, whereas < 1% was positive for the vesicular glutamate transporter (VGluT2). Moreover, of the MCH varicosities, similar to 5% abutted puncta that were immunostained for gephyrin, the postsynaptic AZD1080 nmr marker for GABAergic synapses. In triple-immunostained sections buy GSK461364 viewed with confocal laser scanning microscopy, we established that MCH varicosities that also contained VGAT or abutted upon gephyrin puncta contacted the tyrosine hydroxylase-immunostained neurons of the LC. Our results suggest that although MCH neurons can influence noradrenergic LC neurons through paracrine release and indirect effects
of their peptide, they can also do so through synaptic release and direct postsynaptic effects of GABA and thus serve to inhibit the LC neurons during sleep, when they are silent, and the MCH neurons discharge. (c) 2012 IBRO. Published by Elsevier Ltd. All rights reserved.”
“The product of the human cytomegalovirus (HCMV) UL71 gene is conserved throughout the herpesvirus family. During HCMV infection, protein pUL71 is required for efficient virion egress and is involved in the final steps of secondary envelopment leading to infectious viral particles. We found strong indications for oligomerization of pUL71 under native conditions selleck products when recombinant pUL71 was negatively stained and analyzed
by electron microscopy. Oligomerization of pUL71 during infection was further verified by native and reducing polyacrylamide gel electrophoresis (PAGE). By in silico analyses of the pUL71 sequence, we noticed a basic leucine zipper (bZIP)-like domain, which might serve as an oligomerization domain. We demonstrated the requirement of the bZIP-like domain for pUL71 oligomerization by coimmunoprecipitation and bimolecular fluorescence complementation using a panel of pUL71 mutants. These studies revealed that the mutation of two leucine residues is sufficient to abrogate oligomerization but that intracellular localization of pUL71 was unaffected. To investigate the relevance of the bZIP domain in the viral context, recombinant viruses carrying mutations identical to those in the panel of pUL71 mutants were generated.