| To better understand GABAergic regulation of pituitary cells, and study the function of different GABA-A receptors in physiologically well-characterized systems, I studied molecular composition and pharmacological characteristics of GABA-A receptors in the anterior and neurointermediate lobes of the rat pituitary. Of fourteen GABA-A receptor subunit mRNA species, I developed ribonuclease protection assays for eleven, including the generation of partial cDNA subclones for the {dollar}beta1, beta2, beta3, gamma1,{dollar} and {dollar}delta{dollar} subunits.; Anterior lobe expressed {dollar}alpha1, beta2, beta3{dollar} and {dollar}gamma2{lcub}rm s{rcub}{dollar} mRNAs along with a small amount of {dollar}gamma1{dollar} mRNA; neurointermediate lobe, melanotrophs expressed {dollar}alpha2, alpha3, beta1, beta3, beta1, gamma1{dollar} and {dollar}gamma2{lcub}rm s{rcub}{dollar} mRNAs. The two sites displayed an absolute segregation of {dollar}alpha1{dollar} versus {dollar}alpha2{dollar} and {dollar}alpha3{dollar} expression, and showed beta subunit heterogeneity. The neuro-intermediate lobe expressed as much {dollar}gamma{dollar}1 as {dollar}gamma2{lcub}rm s{rcub}{dollar} mRNA.; Pharmacological properties of pituitary GABA-A receptors were similar to those of brain GABA-A receptors in most respects, including a 2:1 ratio of muscimol to benzodiazepine sites. Quantities of receptor sites in the tissues studied were roughly proportional to the quantities of mRNA detected. As predicted by subunit composition anterior lobe receptors displayed pure Type I benzodiazepine pharmacology while neurointermediate lobe displayed pure Type II pharmacology, allowing clear correlation of these properties with alpha subunit composition in native receptors.; Further, I was able to demonstrate synthesis of 3 alpha-hydroxydihydroprogesterone in the neurointermediate lobe, in what appear to be quantities sufficient to potentiate GABA actions at the GABA-A receptor.; Melanotrophs expressed subunit mRNAs sufficient in theory to direct synthesis of eighteen kinds of receptor with subpopulations possessing differential sensitivity to modulators of the GABA-A receptor. Taken together with the synthesis of a neurally active steroid in the neurointermediate lobe, this suggests a role for endogenous modulators of the GABA-A receptor in controlling actions of melanotrophs. Based on the findings described, I develop a hypothesis on the roles of GABA in melanotroph regulation in the concluding chapter, and present some data on the integration of signals through GABA-A and GABA-B receptors in melanotrophs. |
