A non-inactivating K+ current sensitive to muscarinic receptor activation in rat cultured cerebellar granule neurons

1. Whole-cell recordings were made from cultured cerebellar granule neurons using perforated patch clamp techniques. The primary cultures were prepared using 6- to 9-day-old Sprague-Dawley rats. 2. Neurons in culture for less than 48 h possessed resting membrane potentials of - 29 mV. However, neurons in culture for 7 days had much more hyperpolarized resting membrane potentials (- 89 mV). Over the same period, these neurons developed an additional component of outward current. 3. This non-inactivating current was activated by depolarization, exhibited outward rectification and reversed close to the potassium equilibrium potential. The kinetics of activation and deactivation were very rapid. 4. Muscarine ((+)-muscarine chloride) reversibly inhibited the current with an EC(50) of 0.17 mu M. The inhibition by muscarine was unaffected by pre-incubation for 17-20 h with 120 mu g ml(-1) pertussis toxin. 5. The current and its inhibition by muscarine were unaffected by 100 mu M Cd2+. In Ca2+-free conditions, the current was significantly larger than in 0.5 mM Ca2+, but inhibition by 10 mu M muscarine was significantly reduced. 6. The standing outward current was not obviously affected by 50 mu M 5-HT, 50 mu M noradrenaline, 50 mu M 2-chloroadenosine or 5 mM tetraethylammonium. It was reduced by 10 mu M La3+, 10 mu M Zn2+ and 1 mM Ba2+. 7. Muscarinic agonists increased the input resistance of neurons and shifted the zero current level in the depolarized direction when voltage clamped. This enhanced excitability was evident under current clamp, where 10 mu M muscarine depolarized granule neurons such that action potentials became evident.