Adenovirus-mediated G alpha(q)-protein antisense transfer in neurons replicates G alpha(q) gene knockout strategies

Abstract Antisense approaches are increasingly used to dissect signaling pathways linking cell surface receptors to intracellular effectors. Here we used a recombinant adenovirus to deliver G-protein α q antisense into rat superior cervical ganglion (SCG) neurons and neuronal cell lines to dissect Gα q -mediated signaling pathways in these cells. This approach was compared with other Gα q gene knockdown strategies, namely, antisense plasmid and knockout mice. Infection with adenovirus expressing Gα q antisense (Gα q AS AdV) selectively decreased immunoreactivity for the Gα q protein. Expression of other Gα protein subunits, such as Gα oA/B, was unaltered. Consistent with this, modulation of Ca 2+ currents by the Gα q -coupled M 1 muscarinic receptor was severely impaired in neurons infected with Gα q AS AdV whereas modulation via the Gα oA -coupled M 4 muscarinic receptor was unchanged. In agreement, activation of phospholipase C and consequent mobilization of intracellular Ca 2+ by UTP receptors was lost in NG108-15 cells infected with Gα q AS AdV but not in cells infected with the control GFP-expressing adenovirus. Results obtained with this recombinant AdV strategy qualitatively and quantitatively replicated results obtained using SCG neurons microinjected with Gα q antisense plasmids or SCG neurons from Gα q knockout mice. This combined antisense/recombinant adenoviral approach can therefore be useful for dissecting signal transduction mechanisms in SCG and other neurons.