Activation of metabotropic glutamate receptors induces an outward current which is potentiated by methylxanthines in rat cerebellar Purkinje cells

The responses of slice-cultured Purkinje cells to trans-dl-1-amino-1,3-cyclopentanedicarboxylic acid (t-ACPD) were examined by intracellular recording techniques and fura-2 microfluorometry. Bath-application of t-ACPD (100 μM, 30 s), a selective agonist of metabotropic glutamate receptors (mGluRs), to Purkinje cells voltage-clamped near their resting potential -65 to -60 mV) consistently induced a transient inward current, followed by a slower outward current (I_out). This outward current was characterized by a linear current-voltage relationship in the range from -130 to -60 mV and accompanied by a significant decrease in membrane conductance. The extrapolated reversal potential of I_out was positive to 0 mV. When t-ACPD was applied for 60 s or more it became apparent that I_out emerged in parallel to the wash-out of t-ACPD. Microfluorometric fura-2 measurements in combination with electrophysiological recordings were used to assess the relation between I_out and intracellular free calcium concentration ([Ca²⁺]_i). In contrast to the inward current that was associated with a transient elevation in [Ca²⁺]_i. I_out was not correlated with an elevated [Ca²⁺]_i. When t-ACPD was applied in the presence of caffeine (5 mM), I_out was reversibly enhanced in amplitude. Caffeine affected neither the t-ACPD-induced calcium signal nor the resting [Ca²⁺]_i. While longer applications of caffeine alone induced outward currents with a current-voltage relationship similar to that of I_out, short applications (30 s) of caffeine had no detectable effect per se but still were effective in enhancing I_out when applied in conjunction with t-ACPD. 3-Isobutyl-1-methylxanthine (IBMX, 0.5 mM), a more selective and potent phosphodiesterase inhibitor than caffeine, exhibited caffeine-like effects at a 10-fold lower concentration. We propose that I_out is generated by a transient inhibition of an inward current that is tonically active at rest and largely voltage-independent in the range tested. Our observations provide evidence for an involvement of cyclic nucleotide second messenger systems in the regulation of this current.