This K08 application under the co-mentorship of Drs.  Bob Rosenberg and Gerry Oxford has two goals: 1) to understand the cellular mechanisms that underlie coupling of voltage-activated calcium channels to regulated peptide secretion in the anterior pituitary, and 2) to assess the clinical impact of targeted calcium channel blockade in Congenital Adrenal Hyperplasia (CAH), a disorder resulting from pituitary hypersecretion.  In studies of a pituitary corticotroph cell line (AtT-20) I identified three distinct high voltage-activated calcium channels that are expressed in similar abundance.  Despite this, only the L-type [dihydropyridine (DHP)-sensitive] channel is capable of triggering calcium-dependent secretion of ACTH.  The primary focus of this proposal, therefore, is to examine the mechanisms that underlie the coupling of DHP-sensitive channels to ACTH release.  I hypothesize that differential localization of the L-type channels accounts for the distinct function of the neurosecretory apparatus in ACTH-secreting cells.  I propose to: a) examine the cellular sites of calcium channels and synaptic components using immunocytochemistry/ confocal microscopy, and b) disrupt coupling using a dominant-negative approach.  In CAH, the most common adrenal disorder in children, a molecular defect in cortisol production removes normal negative feedback to the anterior pituitary and results in elevated ACTH secretion.  Elevated ACTH further increases cortisol precursors, and shunts these steroid intermediates to androgen synthetic pathways.  The clinical stigmata (e.g., ambiguous genitalia, virilization and short stature in adults) are due to the excess in androgens.  Treatment requires life-long glucocorticoid replacement to restore the normal feedback mechanism.  Determining the "optimal level" ofglucocorticoid replacement, however, is difficult: under-treatment leads to adrenal insufficiency and androgen excess; over-treatment leads to glucocorticoid excess (e.g., obesity, glucose intolerance and osteopenia).  My work with the pituitary cells in culture suggests that selective pharmacologic blockade of L-type (DHP-sensitive) channels, and, in turn, attenuation of ACTH release, will provide a novel adjunct therapy by allowing lower glucocorticoid dosing.  This would result in enhanced linear growth and improved bone mineral density.  These basic research and clinical studies complement one another, and together, they should enhance our understanding of the cellular mechanisms underlying peptide secretion.  Furthermore, they may allow for improved treatment in hypersecretory conditions, such as CAH.