Dynamin 1 Restrains Vesicular Release to a Subquantal Mode In Mammalian Adrenal Chromaffin Cells

Abstract

Dynamin 1 (dyn1) is required for clathrin-mediated endocytosis in most secretory (neuronal and neuroendocrine) cells. There are two modes of Ca 2ϩ -dependent catecholamine release from single dense-core vesicles: full-quantal (quantal) and subquantal in adrenal chromaffin cells, but their relative occurrences and impacts on total secretion remain unclear. To address this fundamental question in neurotransmission area using both sexes of animals, here we report the following: (1) dyn1-KO increased quantal size (QS, but not vesicle size/content) by Ն250% in dyn1-KO mice; (2) the KO-increased QS was rescued by dyn1 (but not its deficient mutant or dyn2); (3) the ratio of quantal versus subquantal events was increased by KO; (4) following a release event, more protein contents were retained in WT versus KO vesicles; and (5) the fusion pore size (d p ) was increased from Յ9 to Ն9 nm by KO. Therefore, Ca 2ϩ -induced exocytosis is generally a subquantal release in sympathetic adrenal chromaffin cells, implying that neurotransmitter release is generally regulated by dynamin in neuronal cells. Ca 2ϩ -dependent neurotransmitter release from a single vesicle is the primary event in all neurotransmission, including synaptic/ neuroendocrine forms. To determine whether Ca 2ϩ -dependent vesicular neurotransmitter release is "all-or-none" (quantal), we provide compelling evidence that most Ca 2ϩ -induced secretory events occur via the subquantal mode in native adrenal chromaffin cells. This subquantal release mode is promoted by dynamin 1, which is universally required for most secretory cells, including neurons and neuroendocrine cells. The present work with dyn1-KO mice further confirms that Ca 2ϩ -dependent transmitter release is mainly via subquantal mode, suggesting that subquantal release could be also important in other types of cells.

Intracellular injection of phospholipids directly alters exocytosis and the fraction of chemical release in chromaffin cells as measured by nano-electrochemistry

“…2,11 Vesicular content and fraction released have been proposed to play an important role in regulation and plasticity in cellular communication. 2,12,[23][24][25] The fraction of catecholamine released was found to be 74%, 48%, 83%, and 41% for the control, and PC, PE, and LPC treated cells, respectively. These results show that fraction of release for PE is slightly increased compare to the control cells but it is decreased considerably aer intracellular injection of either PC and LPC.…”

Differential Co-release of Two Neurotransmitters from a Vesicle Fusion Pore in Mammalian Adrenal Chromaffin Cells

“…Alternatively, based on the vesicle contents remaining after exocytosis, vesicular secretion can also be classified according to two modes of native transmitter release: ''quantal'' (all-or-none or complete release) and ''sub-quantal'' (''stand-alone foot'' or partial release) . We have previously reported that catecholamine (CA) release tends to be sub-quantal when fusion pore expansion is limited by intrinsic dynamin 1 (Wu et al, 2019) and by either activating an inhibitory G-protein-coupled receptor or knockout (KO) of the Ca 2+ sensor protein synaptotagmin 7 (Syt7) (Schonn et al, 2008;Segovia et al, 2010) in adrenal chromaffin cells. It is known that CA and ATP co-exist in the same dense-core vesicles (DCVs) in chromaffin cells (Esté vez-Herrera et al, 2016;García et al, 2006;Hollins and Ikeda, 1997).…”

Fusion pore expansion and contraction during catecholamine release from endocrine cells

“…The post-spike pore identifies a potential locus for proteins to influence secretion. Many proteins influence spike shape (9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). These results have generally been interpreted as an effect on fusion pore expansion, but these studies lacked a framework for relating spike shape to fusion pores.…”

“…The present analysis provides this framework. Thus, rather than altering pore expansion, the action of dynamin on amperometric spikes (15)(16)(17)19) can be viewed as a change in the size of the post-spike pore or in the speed of the constriction during its formation. This could be related to the collar-forming activity of dynamin around lipidic pores (55).…”

“…The decay of the spike is too slow to be diffusion limited, and dissociation of catecholamine from the proteins of the vesicle matrix has been invoked to account for this feature (3,(6)(7)(8). However, a role for expanding fusion pores is implied by a large number of studies reporting that many fusion proteins influence the spike waveform (9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). Most of these proteins reside in the cytoplasm, out of reach of the matrix within a vesicle.…”