Modulation of α₂-Adrenoceptor Functions by Heterotrimeric Gα_i Protein Isoforms

Abstract

Subtype diversity of heterotrimeric G proteins and G proteincoupled receptors enables a wide spectrum of signal transduction. However, the significance of isoforms within receptor or G protein subfamilies has not been fully elucidated. In the present study, we have tested whether ␣ 2 -adrenoceptors require specific G␣ isoforms for their function in vivo. In particular, we analyzed the role of the highly homologous G␣ i isoforms, G␣ i1 , G␣ i2 , and G␣ i3 , in typical ␣ 2 -adrenoceptor-controlled functions. Mice with targeted deletions in the genes encoding G␣ i1 , G␣ i2 , or G␣ i3 were used to test the effects of ␣ 2 -adrenoceptor stimulation by the agonist medetomidine. The ␣ 2 -adrenoceptor agonist medetomidine inhibited [ 3 H]norepinephrine release from isolated prefrontal brain cortex or cardiac atria tissue specimens with similar potency and efficacy in tissues from wildtype or G␣ i -deficient mice. In vivo, bradycardia, hypotension, induction of sleep, antinociception, and hypothermia induced by ␣ 2 -adrenoceptor activation did not differ between wild-type and G␣ i -knockout mice. However, the effects of the ␣ 2 -agonists medetomidine or 5-bromo-6-(2-imidazolin-2-ylamino)quinoxaline tartrate (UK14,304) on spontaneous locomotor activity or anesthetic sparing were reduced or absent, respectively, in mice lacking G␣ i2 . In microdissected locus coeruleus neurons or postganglionic sympathetic neurons from stellate ganglia, all three G␣ i subunits were expressed as determined by quantitative reverse transcription-polymerase chain reaction, with G␣ i1 and G␣ i2 dominating over G␣ i3 . Functional redundancy of the highly homologous G␣ i isoforms may predominate over specificity to regulate distinct intracellular pathways downstream of ␣ 2 -adrenoceptors in vivo. In contrast, inhibition of locomotor activity and anesthetic sparing may be elicited by a specific coupling of ␣ 2A -adrenoceptors via the G␣ i2 isoform to intracellular pathways.Heterotrimeric G proteins are composed of ␣, ␤, and ␥ subunits and play diverse roles in many aspects of cell regulation. In general, G proteins are classified according to the intracellular signaling pathway stimulated by their ␣ subunits, although growing evidence is supporting a regulatory role for the tightly associated ␤␥ dimers (Gibson and Gilman, 2006;Smrcka, 2008). Agonist activation of G protein-coupled receptors induces a conformational change within the receptor, which subsequently catalyzes the exchange of GDP for GTP on the G␣ subunit leading to the dissociation of the heterotrimer into the GTP-bound G␣ subunit and the functional G␤␥ dimer (Gilman, 1987). These two mediators relay signals from the receptor to several downstream effectors, including ion channels, small GTPases, adenylyl cyclases, phosphodiesterases, and phospholipases, giving rise to the generation of respective second messenger molecules involved in regulating physiological processes (Offermanns, 2003).Based on their sequence homology and differential regulation of effectors, G proteins are g...

Gαi2 Is the Essential Gαi Protein in Immune Complex–Induced Lung Disease

“…In this study, we used mouse strains deficient for Ga i2 or Ga i3 , which have been previously characterized (16,22,24,28). It should be pointed out that Ga i2 -deficient animals were strictly kept under pathogen-free conditions.…”

“…These Ga i knockout (KO) mouse strains were backcrossed to C57BL/6J mice for more than 11 generations and kept in inhalator ventilation cages (IVC) (Ga i2 2/2 ) or under specific pathogen-free (SPF; Ga i3 2/2 ) conditions (24). No obvious signs of intestinal inflammation were visible during the course of the study, and IVC-kept Ga i2 -deficient mice show similar survival as wild-type (WT) mice (see Supplemental Fig.…”

Defective Macrophage Migration in Gαi2- but Not Gαi3-Deficient Mice

“…These Ga i knockout (KO) mouse strains were backcrossed to C57BL/6J (B6) mice for .11 generations and were strictly kept under IVC conditions. Under these conditions, no signs of intestinal inflammation were visible in Ga i2 -deficient mice during the course of the study (15). C5aR-deficient mice backcrossed to C57BL/6J mice for six generations were provided by C. Gerard (Boston, MA) (25).…”

“…Many biological functions of leukocytes, including macrophage phagocytosis and migration, involve Ga i -mediated signaling (11,12). Previous studies in mice lacking Ga i2 or Ga i3 suggested key roles of these two G i isoforms in distinct cellular responses (13)(14)(15)(16). For instance, Ga i3 deficiency leads to an impaired response of insulin-regulated autophagy in liver cells (16,17), whereas Ga i2 2/2 mice show defects in the migration of T and B cells to lymph nodes (18,19) and of eosinophils to sites of allergic tissue injury (20).…”

Repeated exposure to MDMA triggers long-term plasticity of noradrenergic and serotonergic neurons

“…Given that both a 2A -adrenergic and 5-HT 1A receptors are coupled to inhibitory G proteins, 40,41 we used western blotting and qRT-PCR to assess the expression levels of Gi/o-like proteins and mRNAs in the LC, the DRN and the PFC after a repeated MDMA exposure. We found that mice repeatedly injected with MDMA exhibited significant less Gai 1 and Gai 2 immunoreactivity within the LC (t ¼ 3.176, Po0.05 and t ¼ 3.744, Po0.05 for Gai 1 and Gai 2 , respectively) (Figures 5e and f) and Gai 3 immunoreactivity within the DRN (t ¼ 3.042, Po0.05) than controls (Figures 5g and h).…”