Physiological stimulation regulates the exocytic mode through calcium activation of protein kinase C in mouse chromaffin cells

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Abstract
Adrenal medullary chromaffin cells release catecholamines and neuropeptides in an activity-dependent manner controlled by the sympathetic nervous system. Under basal sympathetic tone, catecholamines are preferentially secreted. During acute stress, increased sympathetic firing evokes release of both catecholamines as well as neuropeptides. Both signalling molecules are co-packaged in the same large dense core granules, thus release of neuropeptide transmitters must be regulated after granule fusion with the cell surface. Previous work has indicated this may be achieved through a size-exclusion mechanism whereby, under basal sympathetic firing, the catecholamines are selectively released through a restricted fusion pore, while less-soluble neuropeptides are left behind in the dense core. Only under the elevated firing experienced during the sympathetic stress response do the granules fully collapse to expel catecholamines and neuropeptides. However, mechanistic description and physiological regulation of this process remain to be determined. We employ electrochemical amperometry, fluid-phase dye uptake and electrophysiological capacitance noise analysis to probe the fusion intermediate in mouse chromaffin cells under physiological electrical stimulation. We show that basal firing rates result in the selective release of catecholamines through an Omega-form 'kiss and run' fusion event characterized by a narrow fusion pore. Increased firing raises calcium levels and activates protein kinase C, which then promotes fusion pore dilation until full granule collapse occurs. Our results demonstrate that the transition between 'kiss and run' and 'full collapse' exocytosis serves a vital physiological regulation in neuroendocrine chromaffin cells and help effect a proper acute stress response.