Abstract
1. Effects of cholinergic agonists on the cultured chick cochlear ganglion (CG) neurone were examined using the whole-cell patch-clamp method. 2. Acetylcholine (ACh, 0.1-100 microM) and its non-hydrolysable form, carbamylcholine (CCh, 0.1-300 microM), suppressed the outward current. The CCh-sensitive current was activated at membrane potentials more positive than -70 mV. 3. The CCh-sensitive current slowly activated after step depolarization with a time constant from 20 to 150 ms. The activation time constant decreased monotonically with depolarization of the membrane. 4. The reversal potential of CCh-sensitive current changed as a function of the external K+ concentration (-79, -65 and -44 mV in 5, 10 and 25 mM, respectively) and was approximately equal to the potassium equilibrium potential (-89, -71 and -48 mV in 5, 10 and 25 mM, respectively). The CCh-sensitive current is concluded to be K+ selective. 5. The CCh-sensitive current showed a sigmoid log dose vs. response relationship with an apparent dissociation constant (KD) of 1.4 microM and a Hill coefficient of 1.0. When ACh was applied, an apparent KD of 1.8 microM and a Hill coefficient of 1.0 was measured. 6. The suppression of K+ current by CCh was blocked by atropine (3 microM) and pirenzepine (3 microM), suggesting that the current is mediated by an M1 muscarinic receptor. 7. The CCh suppression of the K+ current was enhanced by GTP-gamma-S (0.1 mM), suggesting that a GTP-binding protein is involved. 8. The CCh suppression of the K+ current was mimicked by protein kinase C activators, 1-oleoyl-2-acetyl-sn-glycerol (OAG, 100 microM), phorbol dibutyrate (PDBu, 2 microM) and phorbol 12-myristate 13-acetate (PMA, 1 microM). The protein kinase inhibitor, staurosporine (0.2 microM) applied internally blocked the CCh suppression of the K+ current which suggests an involvement of protein kinase C.