Strong stimulation triggers full fusion exocytosis and very slow endocytosis of the small dense core granules in carotid glomus cells

Abstract

Chemosensory glomus cells of the carotid bodies release transmitters, including ATP and dopamine mainly via the exocytosis of small dense core granules (SDCGs, vesicular diameter of ∼100 nm). Using carbon-fiber amperometry, we showed previously that with a modest uniform elevation in cytosolic Ca²⁺ concentration ([Ca²⁺]_i of ∼0.5 µM), SDCGs of rat glomus cells predominantly underwent a “kiss-and-run” mode of exocytosis. Here, we examined whether a larger [Ca²⁺]_i rise influenced the mode of exocytosis. Activation of voltage-gated Ca²⁺ channels by a train of voltage-clamped depolarizations which elevated [Ca²⁺]_i to ∼1.6 μM increased the cell membrane capacitance by ∼2.5%. At 30 s after such a stimulus, only 5% of the added membrane was retrieved. Flash photolysis of caged-Ca²⁺ (which elevated [Ca²⁺]_i to ∼16 μM) increased cell membrane capacitance by ∼13%, and only ∼30% of the added membrane was retrieved at 30 s after the UV flash. When exocytosis and endocytosis were monitored using the two-photon excitation and extracellular polar tracer (TEP) imaging of FM1–43 fluorescence in conjunction with photolysis of caged Ca²⁺, almost uniform exocytosis was detected over the cell’s entire surface and it was followed by slow endocytosis. Immunocytochemistry showed that the cytoplasmic densities of dynamin I, II and clathrin (key proteins that mediate endocytosis) in glomus cells were less than half of those in adrenal chromaffin cells, suggesting that a lower expression of endocytotic machinery may underlie the slow endocytosis in glomus cells. An analysis of the relative change in the signals from two fluorescent dyes that simultaneously monitored the addition of vesicular volume and plasma membrane surface area, suggested that with an intense stimulus, SDCGs of glomus cells underwent full fusion without any significant “compound” exocytosis. Therefore, during a severe hypoxic challenge, glomus granules undergo full fusion for a more complete release of transmitters.

Keywords:

• Dense core granules
• type I cells
• chemosensory organ
• dopamine secretion

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

This work was supported by operating grants from the Canadian Institute of Health Research (MOP-79291 to AT, MOP-12070 to FWT) and Natural Science and Engineering Research Council (RGPIN-2016-04653 to FWT); a Visiting Professorship from the University of Tokyo (to FWT), CREST (JPMJCR1652 to HK) from JST; Grant-in-Aid (No. 26221001 to HK), AMED (JP18 dm 01017120 to HK) and World Premier International Research Center Initiative (WPI) to HK from the Ministry of Education, Culture, Sports, Science, and Technology (MEXT) of Japan.