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In the present study, we investigated the effect of ceramide on the outward K[sup +] current in rat pinealocytes using whole cell and single channel recordings. Three components of the whole cell outward K[sup +] current were separated, an iberiotoxin (IBTX)-sensitive K[sup +] current (I[sub KCa]), a transient A current (I[sub A]) and a delayed rectifier current (I[sub K]). C6-ceramide reduced all three components of the outward K[sup +] current. C6-ceramide (30 µm) caused a 53% inhibition of I[sub KCa] [a component that is generated by the IBTX-sensitive K[sup +] channel (BK channel)], a 27% inhibition of I[sub A] and a 17% inhibition of I[sub K]. Additional studies showed that the BK channel was not inhibited by dihydroC6-ceramide, the inactive analog of C6-ceramide, but mimicked by sphingomyelinase which increased intracellular ceramide. The ceramide inhibition of the BK channel was only partly dependent on its inhibition of the L-type Ca[sup 2+] channel. Studies using specific kinase inhibitors showed that calphostin C (a protein kinase C inhibitor) and to a lesser degree lavendustin A (a tyrosine kinase inhibitor) were effective in reducing the ceramide inhibition of I[sub KCa]. Taken together, our results show that, in rat pinealocytes, ceramide reduces the outward K[sup +] current predominantly by inhibiting I[sub KCa]. Moreover, protein kinase C appears to be the main kinase involved in the ceramide inhibition of I[sub KCa]. [ABSTRACT FROM AUTHOR]