cGMP modulation of ACh-stimulated exocytosis in guinea pig antral mucous cells

Abstract

In guinea pig antral mucous cells, ACh stimulates the Ca2+-regulated exocytosis, which has a characteristics feature: an initial transient phase followed by a sustained phase. The effects of cGMP on ACh-stimulated exocytosis were studied in guinea pig antral mucous cells using video microscopy. cGMP enhanced the frequency of ACh-stimulated exocytotic events, whereas cGMP alone did not induce any exocytotic events under the ACh-unstimulated condition. cGMP did not stimulate either Ca2+ mobilization or cAMP accumulation. The Ca2+ dose-response studies demonstrated that cGMP shifted the dose-response curve upward with no shift to the lower concentration. This indicates that cGMP increased maximal responsiveness of the Ca2+-regulated exocytosis, but not the Ca2+ sensitivity. Moreover, under a condition of ATP depletion by dinitrophenol (DNP) or anoxia (N2 bubbling), ACh evoked only a sustained phase in exocytotic events with no initial transient phase. However, ACh evoked an initial transient phase followed by a sustained phase with addition of cGMP before ATP depletion, whereas only a sustained phase was evoked in a case of cGMP addition after ATP depletion. Thus cGMP-induced enhancement in ACh-stimulated exocytotic events requires ATP, suggesting that cGMP modulates ATP-dependent priming of Ca2+-regulated exocytosis in antral mucous cells. In conclusion, cGMP increases the number of primed granules via acceleration of the ATP-dependent priming, which enhances the Ca2+-regulated exocytosis stimulated by ACh.

A PKG inhibitor increases Ca2+-regulated exocytosis in guinea pig antral mucous cells: cAMP accumulation via PDE2A inhibition

“…The previous study demonstrated that 8-BrcGMP enhances the initial phase of Ca 2ϩ -regulated exocytosis in antral mucous cells (14). In this study, we confirmed the same effects of 8-BrcGMP on the frequency of ACh-stimulated exocytotic events.…”

“…An accumulation of cAMP, which accelerates the priming step and the fusion step, also significantly increases the frequency of the initial phase in Ca 2ϩ -regulated exocytotic events. However, cGMP accelerates the priming step of Ca 2ϩ -regulated exocytosis, leading to an increase in the frequency of the initial phase; the cGMP-induced enhancement of the initial phase was not affected by protein kinase A (PKA) inhibitors (PKI-amide and H-89) (14).…”

<b>PPARα induced NOS1 phosphorylation via PI3K/Akt in guinea pig antral mucous cells: NO-enhancement in Ca</b>^<b>2+</b><b>-regulated </b><b>exocytosis </b>

“…In antral mucous cells, the Ca 2+ -regulated exocytosis which is activated by ACh (acetylcholine) is the main mechanism for mucin release (8,14,15,19) and is modulated by many substances, such as cAMP, NO, cGMP, intracellular Cl − , and arachidonic acid (AA) (8,14,19,(21)(22)(23)(24). NO is synthesized by nitric oxide synthase 1 (NOS1) and enhances the Ca 2+ -regulated exocytosis mediated via cGMP accumulation in antral mucous cells (28).…”

“…NO is synthesized by nitric oxide synthase 1 (NOS1) and enhances the Ca 2+ -regulated exocytosis mediated via cGMP accumulation in antral mucous cells (28). Previous studies demonstrated that cGMP increases the releasable pool of mucous granules by accelerating the ATP-dependent priming leading to the enhancement of Ca 2+ -regulated exocytosis in antral mucous cells (11,19). Peroxisome proliferation activation receptor α (PPARα), the natural ligands of which are fatty acids including AA, enhances the Ca 2+ -regulated exocytosis in antral mucous cells (21,28).…”

“…Guinea pigs were sacrificed by intraperitoneal injection of pentobarbital sodium (100 mg/kg), and then, the stomach was removed from the animal. The procedures for the cell preparation have been previously described (8,9,14,15,19,(22)(23)(24). Briefly, the gastric antrum was excised from the stomach, and the mucosal layer was stripped from the muscle layer in cold phosphate buffered saline (4°C) using glass slides.…”