Inhibition of the M current in NG108-15 neuroblastoma × glioma hybrid cells

Abstract

The M current, I _M, a voltage-dependent non-inactivating K current, was recorded in NG108-15 neuroblastoma × glioma hybrid cells, using the whole-cell mode of the patch-clamp technique. We studied inhibition of the M current by bradykinin, phorbol dibutyrate (PDBu), an activator of protein kinase C (PKC), and methylxanthines. Focal application of 0.1–5 μM bradykinin inhibited I _M by about 60%; 5 nM bradykinin inhibited by about 40%. Bath application of 0.1 μM and 1 μM PDBu diminished I _M to about half of the control value. Staurosporine, a PKC inhibitor, applied for 35–43 min in a concentration of 0.3 μM significantly reduced the effect of 1 μM PDBu. M current blockage by PDBu could be partly reversed by bath application of H-7 (51–64 μM), another PKC inhibitor. These observations suggest that the PDBu effect is really due to activation of PKC. The findings are compatible with the view [Brown DA, Higashida H (1988) J Physiol (Lond) 397:185–207] that the bradykinin effect on I _M is mediated by PKC. However, three further observations suggest that this is only true for part of the bradykinin effect. When the suppression of I _M by 1 μM PDBu was fully developed, 0.1 μM bradykinin produced a further inhibition of I _M. Down-regulation of PKC by long-term treatment with PDBu reduced the effect of 0.1 μM bradykinin significantly but did not abolish it. Staurosporine (0.3 μM, applied for 31–46 min) failed to reduce the effect of 5 nM bradykinin significantly. The M current could be reversibly blocked by methylxanthines (caffeine, isobutylmethylxanthine, theophylline) in the millimolar range, probably because of a direct action on the M channels.