Mechanisms of Angiotensin II–Mediated Decreases in Intraneuronal Ca 2+ in Calcium-Loaded Stellate Ganglion Neurons

Abstract

Our laboratory has reported previously that angiotensin II, type-1 (AT 1 ) receptor stimulation in isolated stellate ganglion neurons decreases intraneuronal calcium concentration ([Ca 2+ ]i) acutely if baseline [Ca 2+ ]i is high and increases [Ca 2+ ]i if baseline [Ca 2+ ]i is low. Part of the angiotensin II (Ang II) effect in high Ca 2+ neurons is mediated through stimulation of Na + –Ca 2+ exchange. Current experiments were conducted to identify additional steps in the signaling pathways. In Ca 2+ -loaded neurons, Ang II–induced decreases in [Ca 2+ ]i were attenuated by phospholipase C inhibition (U73122) or nitric oxide (NO) synthase inhibition ( l -NMMA) and were mimicked by the cGMP analogue 8-Br-cGMP. Protein kinase C (PKC) inhibition (bisindolylmaleimide I or Go6976) and protein kinase G (PKG) inhibition (KT5823) partially blocked Ang II–mediated decreases in [Ca 2+ ]i, but complete blockade of Ang II effects was obtained with combined PKC and PKG inhibition. Modulation of inositol triphosphate (IP 3 )-inducible ER Ca 2+ release by [Ca 2+ ]i was investigated using furaptra, an ER-retaining dye. IP 3 -mediated ER Ca 2+ release in β-escin–permeabilized neurons was measured after clamping of [Ca 2+ ]i from 50 nM to 800 nM. Maximal ER Ca 2+ release was observed at ≈200 nM [Ca 2+ ]i, with noted blunting of release at higher [Ca 2+ ]i. Steady-state mRNA transcript and protein levels revealed that the principal IP 3 R isoform expressed was IP 3 R-II. These results suggest that Ca 2+ loading in stellate ganglion neurons promotes Ang II-mediated decreases in [Ca 2+ ]i via PKC and NO/cGMP/PKG pathways and inhibits IP 3 R-II–mediated ER Ca 2+ release.