Heteromeric assembly of GABA(B)R1 and GABA(B)R2 receptor subunits inhibitsCa2+ current in sympathetic neurons

Neuronal GABA(B) receptors regulate calcium and potassium currents via G-pr otein-coupled mechanisms and play a critical role in long-term inhibition o f synaptic transmission in the CNS. Recent studies have demonstrated that a ssembly of GABA(B) receptor GABA(B)R1 and GABA(B)R2 subunits into functiona l heterodimers is required for coupling to potassium channels in heterologo us systems. However whether heterodimerization is required for the coupling of GABA(B) receptors to effector systems in neurons remains to be establis hed. To address this issue, we have studied the coupling of recombinant GAB A(B) receptors to endogenous Ca2+ channels in superior cervical ganglion (S CG) neurons using nuclear microinjection to introduce both sense and antise nse expression constructs. Patch-clamp recording from neurons injected with both GABA(B)R1a/1b and GABA(B)R2 cDNAs or with GABA(B)R2 alone produced ma rked baclofen-mediated inhibition of Ca2+ channel currents via a pertussis toxin-sensitive mechanism. The actions of baclofen were blocked by CGP62349 , a specific GABA(B) antagonist, and were voltage dependent. Interestingly, SCGs were found to express abundantly GABA(B)R1 but not GABA(B)R2 at the p rotein level. To determine whether heterodimerization of GABA(B)R1 and GABA (B)R2 subunits was required for Ca2+ inhibition, the GABA(B)R2 expression c onstruct was microinjected with a GABA(B)R1 antisense construct. This resul ted in a dramatic decrease in the levels of the endogenous GABA(B)R1 protei n and a marked reduction in the inhibitory effects of baclofen on Ca2+ curr ents. Therefore our results suggest that in neurons heteromeric assemblies of GABA(B)R1 and GABA(B)R2 are essential to mediate GABAergic inhibition of Ca2+ channel currents.