Angiotensin II acts intracellularly in vascular smooth muscle cells

Angiotensin (ANG) II is present inside vascular smooth muscle cells (VSMC); however, its intracellular functions if any, are unknown. We tested the hypothesis that intracellular Ang II exerts effects on cytosolic calcium [Ca++]i in VSMC. ANG II was administered via microinjection. Microinjection of ANG II led to a rapid increase in [Ca++]i in the cytosol and in the nucleus. The [Ca++]i increase was due to the influx of extracellular Ca++ ions. The intracellular ANG II effect was totally inhibited by the concomitant injection of the ANG II antagonist CV-11947. Desensitization of extracellular ANG II receptors on the other hand, did not influence the intracellular effects, nor did extracellular CV-11947. The increase in [Ca++]i was not only observed in the microinjected cell, but also in directly adjacent VSMC. In contrast to the microinjected cells, the [Ca++]i increase in the adjacent cells was mostly due to release from intracellular stores. Pretreatment with thapsigargin abolished the ANG II response in adjacent cells. Microinjection of IP3 induced a [Ca++]i response in adjacent cells which was similar to the ANG II-induced effects. Preincubation of VSMC with the uncoupling substances DMSO and heptanol did not decrease the ANG II response, but instead prevented a [Ca++]i surge in adjacent cells. We conclude that intracellular ANG II binds to intracellular ANG II receptors and elicits an increased [Ca++]i in the injected cell and thereafter cells in the immediate neighborhood. Cell-cell contact is necessary for the ANG II-mediated effects. The data suggest that ANG II-related effects may be amplified by an intracellular action.