SEGREGATION OF NITRIC-OXIDE SYNTHASE EXPRESSION AND CALCIUM RESPONSE TO NITRIC-OXIDE IN ADRENERGIC AND NORADRENERGIC BOVINE CHROMAFFIN CELLS

Previous work has demonstrated that nitric oxide can be an important i ntracellular messenger in the regulation of neurosecretion in chromaff in cells. Since standard chromaffin cell cultures are mixed population s of noradrenaline and adrenaline producing cells, it would seem impor tant to understand the functional differences between these individual components. The use of fluorescence imaging techniques for the record ing of cytosolic calcium from single chromaffin cells together with th e immunoidentification of individual cells with specific antibodies ag ainst tyrosine hydroxylase, N-phenyl ethanolamine methyl transferase a nd nitric oxide synthase, has allowed us to measure single-cell calciu m responses in identified adrenergic, noradrenergic and nitrergic chro maffin cells, thus helping us to clarify the differential role of nitr ic oxide in the function of these chromaffin cell types. 53+/-2% of ch romaffin cells were able to synthesize nitric oxide (nitric oxidesynth ase-positive cells), these cells being mainly noradrenergic (82+/-2%). Results indicate that nitric oxide donors such as sodium nitroprussid e, molsidomine and isosorbide dinitrate evoke [Ca2+](i) increases in a 62+/-4% of chromaffin cells, the response to nitric oxide donors bein g between 30 and 50% of that of 20 mu M nicotine. Cells responding to nitric oxide donors were mainly adrenergic (68+/-5%) although 45+/-9% of noradrenergic cells also gave [Ca2+](i) increasing responses. The d istribution of nitric oxide responding cells between nitric oxide synt hase-positive and negative was very similar in the whole population (6 3+/-5 and 60+/-7%, respectively), but these differences were more prom inent when considering the distribution of nitric oxide response betwe en noradrenergic and adrenergic nitric oxide synthase-positive cells; while 73+/-6% of adrenergic nitric oxide synthase-positive cells evoke [Ca2+](i) increases by nitric oxide stimulation, only 35+/-11% of nor adrenergic nitric oxide synthase-positive cells respond. Taken togethe r these results seem to indicate that (i) nitric oxide could act withi n adrenal medulla as both an intracellular and intercellular messenger ; and (ii) noradrenergic cells seem to be specialized in nitric oxide synthesis while adrenergic cells with an endocrine Function could main ly act as a target of neurosecretory action of this second messenger. (C) 1997 IBRO. Published by Elsevier Science Ltd.