Augmentation of L-type calcium current by hypoxia in rabbit carotid body glomus cells: Evidence for a PKC-sensitive pathway

Previous studies have suggested that voltage-gated Ca2+ influx in glomus ce lls plays a critical role in sensory transduction at the carotid body chemo receptors. The purpose of the present study was to determine the effects of hypoxia on the Ca2+ current in glomus cells and to elucidate the underlyin g mechanism( s). Experiments were performed on freshly dissociated glomus c ells from rabbit carotid bodies. Ca2+ current was monitored using the whole cell configuration of the patch-clamp technique, with Ba2+ as the charge c arrier. Hypoxia (pO(2) = 40 mmHg) augmented the Ca2+ current by 24 +/- 3% ( n = 42, at 0 mV) in a voltage-independent manner. This effect was seen in a CO2/HCO3--, but not in a HEPES-buffered extracellular solution at pH 7.4 ( n = 6). When the pH of a HEPES-buffered extracellular solution was lowered from 7.4 to 7.0, hypoxia augmented the Ca2+ current by 20 +/- 5% (n = 4, at 0 mV). Nisoldipine, an L-type Ca2+ channel blocker (2 mu M, n = 6), preven ted, whereas, omega-conotoxin MVIIC (2 mu M, n = 6), an inhibitor of N and P/Q type Ca2+ channels, did not prevent augmentation of the Ca2+ current by hypoxia, implying that low oxygen affects L-type Ca2+ channels in glomus c ells. Protein kinase C (PKC) inhibitors, staurosporine (100 nM, n = 6) and bisindolylmaleimide (2 mu M, n = 8, at 0 mV), prevented, whereas, a protein kinase A inhibitor (4 nM PKAi, n = 10) did not prevent the hypoxia-induced increase of the Ca2+ current. Phorbol 12-myristate 13-acetate (PMA, 100 nM ), a PKC activator, augmented the Ca2+ current by 20 6 3% (n = 8, at 0 mV). In glomus cells treated with PMA overnight (100 nM), hypoxia did not augme nt the Ca2+ current (-3 + 4%, n = 5, at 0 mV). Immunocytochemical analysis revealed PKC delta-like immunoreactivity in the cytosol of the glomus cells . Following hypoxia (6% O-2 for 5 min), PKC delta-like immunoreactivity tra nslocated to the plasma membrane in 87 6 3% of the cells, indicating PKC ac tivation. These results demonstrate that hypoxia augments Ca2+ current thro ugh L-type Ca2+ channels via a PKC-sensitive mechanism.