PATCH-CLAMP STUDY OF POSTNATAL-DEVELOPMENT OF CA1 NEURONS IN RAT HIPPOCAMPAL SLICES - MEMBRANE EXCITABILITY AND K+ CURRENTS

1. The postnatal development of membrane properties and outward K+ currents in CA1 neurons in rat hippocampal slices was studied with the use of whole-cell patch-clamp techniques. 2. Neurons at all postnatal ages (2-30 days; P2-30) were capable of generating tetrodotoxin (TTX)-sensitive action potentials in response to intracellular injection of depolarizing current pulses. There was gradual increase in the amplitude and a decrease in the duration of these action potentials with age. Stable values for spike duration were reached by P15, whereas spike amplitude increased until P20-25. In P2-5 neurons, the duration of action potentials was greatly prolonged by depolarization from the resting membrane potential, indicating a weak spike repolarizing mechanism at depolarized potentials. In contrast, the duration of spikes evoked in P20-30 neurons was not affected by similar changes in the membrane potential. 3. Application of tetraethylammonium (TEA, 10mM) had no effect on the duration of spikes in P3-5 neurons, whereas application of 4-aminopyridine (4-AP, 2 mM) produced large increases in spike duration. In contrast, the duration of spikes in P26 neurons was greatly increased after TEA application, whereas 4-AP had smaller effects on spike duration in these neurons. 4. The input resistance and membrane time constant decreased with age from P2 to P15. The values for both parameters were considerably greater than those reported with conventional intracellular recording electrodes in the immature hippocampus. The resting membrane potentialf became more hyperpolarized with age. When the recording pipettes contained KCl (140 mM), the resting potential of P3-4 neurons was 34 mV depolarized compared with resting potentials observed with potassium gluconate-filled pipettes. Only a 13-mV change in resting potential was observed during similar comparisons in P27-28 neurons. 5. Outward currents activated by depolarization were examined with the use of voltage-clamp techniques in P2-30 neurons. In P2-5 cells, a small, slowly inactivating outward current was evoked with depolarizing commands from holding potentials near -50 mV. By preceding the depolarizing commands with a hyperpolarizing prepulse, an additional early transient outward current was evoked. The sustained and transient outward currents were separated by their kinetic properties and their sensitivity to cobalt (Co2+), TEA, and 4-AP. 6. The transient outward current (I(A)) was the most prominent current in P2-5 neurons. The amplitude of I(A) was reduced by Co2+ without affecting the rate of inactivation. The 4-AP (2 mM) reduced the amplitude and increase the rate of inactivation of I(A), whereas TEA (10 mM) had no effect. In P30 neurons, depolarizing commands evoked large TEA- and Co2+ -sensitive outward currents that overlapped I(A). It is suggested that I(A) is the predominant outward current underlying spike repolarization in P2-5 neurons. The TEA-sensitive sustained currents begin to play a greater role in spike repolarization after the first week of postnatal development. 7. The sustained outward currents were blocked by TEA (10 mM) and were little affected by 4-AP (2 mM) applications. Comparison of the Co2+ -insensitive sustained current between P3-5 and P25-30 neurons showed a great increase in its amplitude with age. A calcium-dependent component of outward currents was observed in neurons of all ages. However, the sustained Ca2+ -dependent currents only became prominent after the first week of postnatal development. 8. Our results indicate that considerable development of excitable membrane properties occurs during the first 2 wk of postnatal development in CA1 hippocampal neurons. The results are consistent with an overall increase in the number of voltage-dependent ionic channels during development, especially those underlying TEA-sensitive K+ currents.