Nitric oxide modulation of calcium‐activated potassium channels in postganglionic neurones of avian cultured ciliary ganglia

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Abstract
1 A study has been made of the modulation of calcium-activated potassium channels in cultured neurones of avian ciliary ganglia by sodium nitroprusside and l-arginine. 2 Sodium nitroprusside (100 μm) reduced the net outward current by 22 ± 1% at 4.8 ms (mean ± s.e.mean) and 25 ± 1% at 350 ms during a test depolarization to + 40 mV from a holding potential of −40 mV. The outward current remained reduced for the duration of the recording following a single application of sodium nitroprusside. These effects did not occur if the influx of calcium ions was first blocked with Cd2+ (500 μm). Application of ferrocyanide (100 μm) reduced the net outward current by only 6 ± 3% at 350 ms during a test depolarization to + 40 mV. 3 l-Arginine (270 μm) reduced the net outward current on average by 19 ± 2% at 4.8 ms and 22 ± 2% at 350 ms during a test depolarization to + 40 mV. The current remained in this reduced state for the duration of the recording following a single application of l-arginine. These effects were reduced to 11 ± 1% at 4.8 ms and 11 ± 2% at 350 ms in the presence of Nω-nitro-l-arginine methyl ester (l-NAME, 100 μm). 4 In order to alleviate the dependence of calcium-activated potassium channels (Ik(Ca)) on the inward flux of calcium ions, the patch-clamp pipettes were filled with a solution containing 100 μm CaCl2, and the Ca2+ in the bathing solution was replaced with EGTA. Under these conditions sodium nitroprusside reduced the total outward current during a depolarizing pulse of + 40 mV by 9 ± 1 % at 4.8 ms and by 36 ± 3% at 350 ms. l-Arginine (270 μm) reduced this current under the same conditions by 9 ± 1% at 4.8 ms and by 35 ± 2% at 350 ms. 5 Calcium-activated potassium currents were sensitive to apamin (50 nm), as this reduced the outward current by 23 ± 3% at 350 ms when a high calcium-containing pipette was used during a depolarizing command to + 40 mV. l-Arginine still decreased the outward current in the presence of apamin (50 nm), by 5 ± 1% at 4.8 ms and by 19 ± 2% at 350 ms, indicating that l-arginine could reduce an apamin-insensitive Ik(Ca). 6 Calcium-activated potassium currents were also sensitive to charybdotoxin (10 nm), as this reduced the outward current by 34 ± 4% at 350 ms when a high calcium-containing pipette was used during a depolarizing command to + 40 mV. l-Arginine still decreased the outward current in the presence of charybdotoxin, by 6 ± 1% at 4.8 ms and 12 ± 4% at 350 ms, showing that l-arginine could reduce a charybdotoxin-insensitive Ik(Ca). 7 The present results indicate that NO-synthase in ciliary ganglia can modulate Ik(Ca) by a method which is independent of the action of NO on the calcium channels. The Ik(Ca) is decreased significantly at 4.8 ms into a depolarizing pulse, at a time that would decrease the rate of repolarization of the action potential. Ik(Ca) is also reduced at longer times (350 ms), indicating an affect on the inactivating process.