Journal of neurochemistry
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Journal of neurochemistry · Jun 2010
Nuclear localization of the G protein beta 5/R7-regulator of G protein signaling protein complex is dependent on R7 binding protein.
The neuronally expressed G beta(5) subunit is the most structurally divergent among heterotrimeric G beta isoforms and unique in its ability to heterodimerize with the R7 subfamily of regulator of G protein signaling (RGS) proteins. The complex between G beta(5) and R7-type RGS proteins targets the cell nucleus by an unknown mechanism. Although the nuclear targeting of the G beta(5)/R7-RGS complex is proposed to involve the binding of R7-binding protein (R7BP), this theory is challenged by the observations that endogenous R7BP is palmitoylated, co-localizes strongly with the plasma membrane, and has never been identified in the cytosol or nucleus of native neurons or untreated cultured cells. ⋯ As the Disheveled, EGL-10, Pleckstrin homology domain is essential for R7BP binding to RGS7, we studied the subcellular localization of G beta(5) in primary neurons and brain from mice deficient in R7BP. The level of endogenous nuclear G beta(5) and RGS7 in neurons and brains from R7BP knockout mice is reduced by 50-70%. These results suggest that R7BP contributes significantly to the nuclear localization of endogenous G beta(5)/R7-RGS complex in brain.
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Journal of neurochemistry · Jun 2010
Regional differences in nerve terminal Na+ channel subtype expression and Na+ channel-dependent glutamate and GABA release in rat CNS.
We tested the hypothesis that expression of pre-synaptic voltage-gated sodium channel (Na(v)) subtypes coupled to neurotransmitter release differs between transmitter types and CNS regions in a nerve terminal-specific manner. Na(v) coupling to transmitter release was determined by measuring the sensitivity of 4-aminopyridine (4AP)-evoked [(3)H]glutamate and [(14)C]GABA release to the specific Na(v) blocker tetrodotoxin (TTX) for nerve terminals isolated from rat cerebral cortex, hippocampus, striatum and spinal cord. Expression of various Na(v) subtypes was measured by immunoblotting using subtype-specific antibodies. ⋯ The relative nerve terminal expression of total Na(v) subtypes as well as of specific subtypes varied considerably between CNS regions. The region-specific potencies of TTX for inhibition of 4AP-evoked glutamate release correlated with greater relative expression of total nerve terminal Na(v) and Na(v)1.2. Nerve terminal-specific differences in the expression of specific Na(v) subtypes contribute to transmitter-specific and regional differences in pharmacological sensitivities of transmitter release.
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Journal of neurochemistry · Jun 2010
Neurochemical properties of enkephalinergic neurons in lumbar spinal dorsal horn revealed by preproenkephalin-green fluorescent protein transgenic mice.
Enkephalin (ENK) has been implicated in nociceptive transmission in the spinal cord while its functional role is not clear because of difficulties in ideally visualizing ENKergic neurons. We thus developed preproenkephalin-green fluorescent protein transgenic mice to better identify ENKergic neurons. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) together with immunohistochemistry and in situ hybridization were first employed to confirm the successful transgenic manipulation and its application in showing spinal ENKergic neurons. ⋯ GABA was found to exist in 42.9 +/- 2.8% of ENKergic neurons that were mainly located in lamina I-III. The proportions of parvalbumin-, calretinin-, calbindin- and neuronal nitric oxide synthase-positive cells among the ENKergic neurons were 5.2 +/- 0.7%, 42.6 +/- 2.3%, 25.8 +/- 2.2% and 11.1 +/- 1.6%, respectively. Compared with previously findings obtained with ENK antibody labeling, this line of newly generated mice can be a reliable tool for the study of specific spinal ENKergic neuronal population.