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Activator of G protein Signaling 1 (AGS 1) and Ras homologue enriched in striatum (Rhes) define a new group of Ras-like monomeric G proteins whose signaling properties and physiological roles are just beginning to be understood. Previous results suggest that AGS1 and Rhes exhibit distinct preferences for heterotrimeric G proteins, with AGS 1 selectively influencing G[alpha]i and Rhes selectively influencing Gas. Here, we demonstrate that AGS1 and Rhes trigger nearly identical modulation of N-type [Ca.sup.2+] channels ([Ca.sub.v]2.2) by selectively altering G[alpha]i-dependent signaling. Whole-cell currents were recorded from HEK293 cells expressing [Ca.sub.v]2.2 and G[alpha]i- or G[alpha]s-coupled receptors. AGS1 and Rhes reduced basal current densities and triggered tonic voltage-dependent (VD) inhibition of [Ca.sub.v]2.2. Additionally, each protein attenuated agonist-initiated channel inhibition through God-coupled receptors without reducing channel inhibition through a Gas-coupled receptor. The above effects of AGS1 and Rhes were blocked by pertussis toxin (PTX) or by expression of a G[beta][gamma]-sequestering peptide (masGRK3ct). Transfection with HRas, KRas2, Rap1A-G12V, Rap2B, Rheb2, or Gem failed to duplicate the effects of AGS1 and Rhes on [Ca.sub.v]2.2. Our data provide the first demonstration that AGS1 and Rhes exhibit similar if not identical signaling properties since both trigger tonic G[beta][gamma] signaling and both attenuate receptor-initiated signaling by the G[beta][gamma] subunits of PTX-sensitive G proteins. These results are consistent with the possibility that AGS1 and Rhes modulate [Ca.sup.2+] influx through [Ca.sub.v]2.2 channels under more physiological conditions and thereby influence [Ca.sup.2+]-dependent events such as neurosecretion. voltage-gated calcium channels; Ras-like; Dexras 1; Dexras2; RASD 1; RASD2

[Ca.sup.2+] influx through voltagegated R-type ([Ca.sub.v]2.3) [Ca.sup.2+] channels is important for hormone and neurotransmitter secretion and other cellular events. Previous studies have shown that [Ca.sub.v]2.3 is both inhibited and stimulated through signaling mechanisms coupled to muscarinic ACh receptors. We previously demonstrated that muscarinic stimulation of [Ca.sub.v]2.3 is blocked by regulator of G protein signaling (RGS) 2. Here we investigated whether muscarinic inhibition of [Ca.sub.v]2.3 is antagonized by RGS3. RGS3 is particularly interesting because it contains a lengthy (~380 residue) amino-terminal domain of uncertain physiological function. [Ca.sub.v]2.3, [M.sub.2] muscarinic ACh receptors ([M.sub.2]R), and various deletion mutants of RGS3, including its native isoform RGS3T, were expressed in HEK293 cells, and agonist-dependent inhibition of [Ca.sub.v]2.3 was quantified using whole cell patch-clamp recordings. Full-length RGS3, RGS3T, and the core domain of RGS3 were equally effective in antagonizing inhibition of [Ca.sub.v]2.3 through [M.sub.2]R. These results identify RGS3 and RGS3T as potential physiological regulators of R-type [Ca.sup.2+] channels. Furthermore, they suggest that the signaling activity of RGS3 is unaffected by its extended amino-terminal domain. Confocal microscopy was used to examine the intracellular locations of four RGS3-enhanced green fluorescent protein fusion proteins. The RGS3 core domain was uniformly distributed throughout both cytoplasm and nucleus. By contrast, full-length RGS3, RGS3T, and the amino-terminal domain of RGS3 were restricted to the cytoplasm. These observations suggest that the amino terminus of RGS3 may serve to confine it to the cytoplasmic compartment where it can interact with cell surface receptors, heterotrimeric G proteins, and other signaling proteins. calcium channels; regulator of G protein signaling proteins; muscarinic acetylcholine receptors; enhanced green fluorescent protein-fusion proteins; voltage-gated R-type calcium channels

The structural regions of the cardiac channel alpha sub(1C) subunit involved in calcium-dependent inactivation was probed through analyzing a series of chimeric alpha1 subunits wherein the alpha sub(1S) replaced the major cytoplasmic domains of alphasub(1) channels. Results have indicated that the cytoplasmic carboxyl terminus and the I-II/II-III loops mediated the molecular mechanism of calcium-dependent inactivation.

The investigation of the ability of BI (class A) Ca2+ channels to mediate excitation-contraction (E-C) coupling in dysgenic skeletal muscles reveals the presence of heterologous expression in these channels. The investigation is performed by injecting expression plasmids with cDNA encoding BI channels into dysgenic mouse myotubule nuclei. The whole-cell patch-clamp method records the intermembrane charge movement and ionic currents produced by the BI channels. The density of ionic BI Ca2+ channel current is very high in the myotubules. Spontaneous contractions are not displayed by the myotubules. The average maximal charge movement is low in the BI channels while the average maximal conductance is very large.

The dihydropyridine (DHP) receptor in skeletal muscle serves both as a calcium channel and as a voltage sensor for excitation-contraction coupling. These two functions have been proposed to correspond to the two different forms of DHP receptors, the long and the short. But the short form performs both functions, as revealed by the construction of a protein corresponding to the truncated DHP receptor, using a complimentary DNA. It has not been determined whether these two functions are performed simultaneously by individual DHP receptors.