Based on previous work, the investigator proposes that a reduction in GABA tone is the critical change initiating puberty in rhesus monkeys and that this leads to subsequent establishment of facilitatory neuronal pathways. The first three Specific Aims will further test this hypothesis and examine possible molecular mechanisms underlying the change in GABA tone. The experiments in Specific Aim 1 will determine if blockade of GABA action leads to an increase in glutamate release in the median eminence and in the response to NMDA ,and monitor changes in glutamic acid decarboxylase (GAD) during puberty. GAD mRNA and protein levels will be measured using ribonuclease protection assay, in situ hybridization (ISH), and immunocytochemistry (ICC), and GAD bioactivity measured in vitro.
In Specific Aim 2, similar techniques will be use to examine expression of the GABA transporter (GAT-1) during puberty, and the importance of GAT-1 to control of LHRH will be tested by administration of an antagonist or antisense DNA.
In Aim 3, they will examine the possibility that changes in the subunit composition of the GABA receptor occur in the hypothalamus and, more specifically, in LHRH neurons using dual ICC an ISH.
Specific Aim 4 will test the hypothesis that a decrease in tonic inhibition by neuropeptide-Y (NPY) contributes to puberty by monitoring NPY mRNA and protein levels, determining if antisense DNA to NPY alters LHRH, GABA, or the response of LHRH neurons to NMDA, and if antisense DNA to GAD increases NPY.
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