Development of Voltage-Activated Potassium Currents in Cultured Cerebellar Granule Neurons Under Different Growth Conditions. Gorter, J. A., E. Aronica, N. J. Hack, R. Balzs, and W. J. Wadman. Experimental Zoology, University of Amsterdam, 1098 SM Amsterdam; and Netherlands Institute for Brain Research, Amsterdam, The Netherlands.
APStracts 2:0015N, 1995.
SUMMARY AND CONCLUSIONS
1 . The functional expression of two potassium currents in cultured cerebellar granule cells was investigated with the whole cell patch-clamp technique in relation to development and growth condition. Cells were grown in medium containing different concentrations of potassium: 25 mM (K25) and 40 mM (K40), together referred to as "high K + " 10 mM (K10) or "low K + " ; and K10 with 100 [mu]M N -methyl-d-aspartate (KNMDA). All conditions are known to influence maturation and survival of granule cells in culture. 2 . At 2 days in vitro (DIV) the membrane capacitance, taken as index of membrane surface area, was the same for cells grown in each growth condition. At 7-9 DIV it had increased in each condition, but to a substantially larger extent in cells grown in KNMDA, K25, and K40 than in cells grown in K10. During development the input resistance only decreased in cells grown in KNMDA and high K + . 3 . A delayed potassium current ( I K ) and a fast transient potassium current (I A ) could both be recorded at 2 DIV in each growth condition, although a few neurons only expressed the I K . The I K was partially suppressed by tetraethylammonium (5 mM), whereas I A was predominantly sensitive to 4- aminopyridine (5 mM). 4 . Normalized for cell capacitance, the specific I A conductance hardly changed during development in cells grown in high K + and KNMDA. Cells in K10, however, displayed an I A with totally different properties in 23 of 24 cells; the specific I A conductance in these cells was considerably smaller at 7-9 DIV, suggesting a deletion of these channels during development. 5 . At 2 DIV, I A inactivated monoexponentially with a time constant that did not significantly differ between growth conditions. Thetime constant was slightly voltage dependent. At 7-9 DIV the voltage dependence became more prominent, in particular at voltages more positive than+10 mV. In many cells grown in K10 in which a transient current still could be detected, the voltage dependence was even stronger, with the fastest inactivation at -0 mV. 6 . The voltage of half- maximal steady-state inactivation of I A was 90.8 +/- 0.3 (SE) mV at 2 DIV for cells in each growth condition. It shifted to more depolarized levels at 7-9 DIV, dependent on growth condition: 80 mV in K10 and KNMDA, 77 mV in K25, and 72 mV in K40. At 2 DIV the slope factor of the Boltzmann function describing the voltage-dependent removal of inactivation was 8.2 +/- 0.9 mV, which did not change at 7-9 DIV, when it was 8.9, 8.4, and 8.0 mV in K10, K25, and K40, respectively. 7 . The activation of the I K was voltage dependent and half- maximal at 0 mV in all growth conditions and independent of time in culture. During development the specific I K conductance increased in the cells grown in the K10 condition but not in cells grown in the other conditions. Because I A was greatly reduced in this condition, there seems to be a reciprocal modulation of these two currents in neurons grown in K10. 8 . These findings suggest that the functional expression of I A and I K in cerebellar granule cells in culture can be regulated by activity-dependent processes during a critical period of development. Because the high K + and KNMDA conditions could mimic the early afferentiation of granule cells, similar mechanisms may play a role in the voltage gated potassium channel expression in cerebellar granule cells in situ. 

Received 8 June 1994; accepted in final form 6 January 1995.
APS Manuscript Number J280-3.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on  3 April 1995.