The GTP-binding protein, Go9 regulates neuronal calcium channels

Abstract

In neuronal cells, opioid peptides and opiates inhibit neurotransmitter release, which is a calcium-dependent process. They also inhibit adenylyl cyclase, presumably via the membrane signal-transducing component, Gi, a guanine nucleotide-binding protein (G-protein). No causal relationship between these two events has yet been demonstrated. Besides Gi, membranes of neuronal tissues contain large amounts of Go, a G-protein with unknown function. Both G-proteins are heterotrimers consisting of alpha-, beta- and gamma-subunits; the alpha-subunits can be ADP-ribosylated by an exotoxin from Bordetella pertussis (PT), which modification inhibits receptor-mediated activation of the G-protein. It was recently shown that noradrenaline, dopamine and gamma-aminobutyric acid (GABA) inhibit the voltage-dependent calcium channels in dorsal root and sympathetic ganglia; this inhibition is mimicked by intracellular application of guanine nucleotides and blocked by PT, suggesting the involvement of a G-protein. Here we report an inhibitory effect of the opioid D-Ala2, D-Leu5-enkephalin (DADLE) on the calcium current (ICa) in neuroblastoma X glioma hybrid cells (N X G cells). Pretreatment with PT almost completely abolishes the DADLE effect. The effect is restored by intracellular application of Gi and Go. As the alpha-subunit of Go (with or without beta-gamma complex) is 10 times more potent than Gi, we propose that Go is involved in the functional coupling of opiate receptors to neuronal voltage-dependent calcium channels.

G proteins as regulators of ion channel function

“…Pertussis toxin (PTX, a bacterial exotoxin that blocks the receptor-mediated activation of certain G proteins, notably G i and G o , through the ADP ribosylation of the α-subunit) interferes with the norepinephrine (NE) and GABA receptormediated inhibition of Ca 2+ currents recorded from dorsal root ganglion (DRG) neurons 16 . Similarly, PTX also blocks the inhibitory actions of the enkephalin analog D-Ala-D-Leuenkephalin (DADLE) and somatostatin on Ca 2+ currents recorded from the neuroblastomaglioma cell line NG108-15, and the pituitary cell line AtT20, respectively 13,17 . As mentioned above, activation of G proteins is terminated by hydrolysis of the bound GTP.…”

“…As mentioned above, activation of G proteins is terminated by hydrolysis of the bound GTP. Intracellular application of the non-hydrolysable GTP analog, GTP-γ-S, converts the normally reversible responses to agonist in these cells into irreversible responses 13,17,18 . In contrast, intracellular application of the GDP analog GDP-β-S (which competes with GTP for the nucleotide binding site) blocks the inhibitory actions of NE and DADLE on Ca 2+ currents recorded from DRG and NG108-15 cells, respectively 13,16 .…”

“…Similarly, it has been thought that G o , a regulatory protein found in abundance in neural tissue (~1% of total protein), might also be linked to enzyme effectors other than adenylate cyclase but, until recently, the α-subunit of G o has had no demonstrable physiological function. A recent paper by Hescheler et al 13 indicates that α o may play a pivotal role in regulating Ca 2+ channel function in neurons. Furthermore, the recently reported co-localization of α o immunoreactivity and phorbol ester binding in brain 14 suggests the possibility that physiological processes regulated by this G protein may be mediated by protein kinase C.…”

Excitotoxin lesion of nucleus basalis causes a specific decrease in Go mRNA in cerebral cortex

“…A similar effect can be demonstrated with opioid peptides in neuroblastoma cells where it has been specifically shown that the pertussis toxin block can be prevented by intracellular application of Gi and G,, the latter effect of G, being 10 times more potent than Gi [17]. This is further supported from the evidence that rat brain G, is a substrate of pertussis toxin ADPribosylation [ 181.…”

The role of G proteins in transmembrane signalling

“…For other effectors the situation is even less clear: ac with GTPyS bound activates Ca2+ channels (Yatani et al, 1987b), but other ac subunits are also effective (Hescheler et al, 1987). The G proteins that regulate polyphosphoinositide-specific phospholipase C are particularly elusive, with evidence for regulation by both pertussis toxin-sensitive and -insensitive G proteins (Taylor & Merritt, 1986;Harden, 1989).…”