Regulation of cytoplasmic calcium levels by two nitric oxide receptors

We examined the effects of dissolved nitric oxide (NO) gas on cytoplasmic calcium levels ([Ca²⁺]ⁱ) in C6 glioma cells under anoxic conditions. The maximum elevation (27 ± 3 nM) of [Ca²⁺]ⁱ was reached at 10 μM NO. A second application of NO was ineffective if the first was >0.5 μM. The NO donor diethylamine/NO mimicked the effects of NO. Acute exposure of the cells to low calcium levels was without effect on the NO-evoked response. Thapsigargin (TG) increased [Ca²⁺]_i and was less effective if cells were pretreated with NO. Hemoglobin inhibited the effects of NO at a molar ratio of 10:1. 8-Bromo-cGMP was without effect on the NO-evoked response. If cells were pretreated with TG or exposed chronically to nominal amounts of calcium, NO decreased [Ca²⁺]_i. The results suggest that C6 glioma cells have two receptors for NO. One receptor (NOA) elevates [Ca²⁺]ⁱ and resides on the endoplasmic reticulum (ER). The other receptor (NO_B) decreases [Ca²⁺]ⁱ and resides on the plasmalemma or the ER. The latter receptor dominates when the level of calcium within intracellular stores is diminished.