Myr+-Gi2α and Goα subunits restore the efficacy of opioids, clonidine and neurotensin giving rise to antinociception in G-protein knock-down mice ☆

Abstract In mice whose Gi/o-protein function had been impaired by antisense ‘knock-down’ or pertussis toxin treatment, icv injection of myr + -G i/o α subunits restored the effectiveness of β-endorphin, morphine, DPDPE, clonidine and neurotensin to produce antinociception. Myr + -Gα subunits of the class of G-proteins actually impaired were more effective than unlike but related myr + -Gα subunits. Selectivity was noted in that only exogenous myr + -Gα subunits affected (enhanced) the activity of agonists in Gα-deficient signalling systems. This treatment had little effect on agonist potency when the impairment resided at the receptor level. The potential of the opioids, clonidine and R-PIA to increase Gα-related in vitro hydrolysis of GTP was also re-established after injecting myr + -G i2 α subunits into Gi2-knocked-down mice. Myr + -G i2 α subunits pre-incubated with GTPγS or GDPβS before icv injection did not improve the activity of agonists in vivo (antinociception) or in vitro (regulation of low K m GTPase). After impairing the function of PKCβ1 by antisense treatment or with the inhibitor H7, the effect of myr + -Gα subunits on agonist potency was prevented. Electron microscope analysis showed the entry of gold-conjugated myr + -Gα subunits into neural cells. These particles were found in the cytoplasm, associated with the plasma membranes of different neuronal processes and also in synaptic junctions. In cultured neurons and astrocytes myr + -G i2 α-associated fluorescence was internalised in a dose-dependent manner and distributed in the plasma membrane and cytosol, as well as in nuclei of dividing astrocytes. Thus, Gα subunits in CSF enter into neurons and functionally couple to the receptor-triggered signalling cascade. As G-proteins have been implicated in the pathophysiology of several neural disorders, this finding may be valuable in the therapy of such dysfunctions.