Upregulation of the hyperpolarization-activated cation current in rat thalamic relay neurones by acetazolamide

1. The effect of inhibition of brain carbonic anhydrase (CB) on the hyperpolarization-activated cation current (I-h) of thalamocortical (TC) neurones of the rat ventrobasal thalamic complex (VB) was investigated in an in vitro slice preparation using the whole-cell patch-clamp technique and fluorescence ratio imaging of the pH indicator 2',7'-bis(carboxyethyl) -5(and -6)-carboxyfluorescein (BCECF). 2. Recording of I-h before and after addition of 0.4-0.8 mM acetazolamide to the bathing fluid revealed a significant shift in the voltage dependence of activation (V-1/2) of 5-7 mV to more positive potentials. 3. Simultaneous recording of I-h and BCECF fluorescence ratio (F-420/F-495) revealed an increase in I-h amplitude accompanied by an intracellular alkalinization upon application of acetazolamide. The CA inhibitor ethoxyzolamide (EZA, 50 mu M) also led to an intracellular alkalinization and a subsequent 4-5 mV positive shift of V-1/2 of I-h. 4. Acetazolamide and EZA both profoundly slowed the rapid fall of pH, upon switching from Hepes- to CO2/HCO3--buffered superfusate, indicating intracellular CA isoforms in TC neurones. 5. In slices bathed in Hepes-buffered saline, addition of acetazolamide had no effect on the amplitude and time course of activation of I-h, indicating that the action of acetazolamide on I-h was dependent on the presence of HCO3-. 6. Under current-clamp conditions, the neuronal response to hyperpolarizing current pulses in the presence of acetazolamide was decreased as compared to control. This resulted in a strongly reduced ability of TC neurones to produce rebound Ca2+-mediated spikes. 7. The present results implied that in TC neurones acetazolamide led to an intracellular alkalinization which causes, due to its pH sensitivity, an increase in I-h.