Chimeric G proteins made by replacing the C-terminal heptapeptide of Gq with the C-terminal heptapeptide of Gs or Gi were used to assess the relative coupling of 3-adrenergic receptor (₃-AR) splice variants (_3A & _3B) to Gs and Gi. The Gq/s and Gq/i chimeras transformed the response to receptor activation from regulation of adenylylcyclase to mobilization of intracellular calcium [Ca²⁺]i. Complementary high throughput and single cell approaches were used to evaluate agonist-induced coupling of the receptor to the G protein chimeras. In cells stably transformed with rat ₃-AR and transfected with the G protein chimeras, ₃-AR-induced coupling to Gq/s produced a rapid 8-fold increase in [Ca²⁺]i followed by a slow decay to levels 25% above baseline. Gq/i also linked rat ₃-AR to mobilization of [Ca²⁺]i, but the net 2.5 fold increase in [Ca²⁺]i was only 30% of the response obtained with Gq/s. Activation of the rat ₃-AR also increased GTP binding to endogenous Gi by 3-fold in membranes from CHO cells stably transformed with receptor. A complementary single cell imaging approach was used to assess the relative coupling of mouse _3A- and _3B-AR to Gi. The _3A- and _3B-AR coupled equivalently to Gq/i but the temporal patterns of [Ca²⁺]i mobilization indicated that coupling was significantly less efficient than coupling to Gq/s. Collectively, these findings indicate less efficient but equivalent coupling of _3A- and _3B-AR to Gi vs Gs, and suggest that differential expression of the splice variants would not produce local differences in signaling networks linked to ₃-AR activation.