Genetic alteration of the α2-adrenoceptor subtype c in mice affects the development of behavioral despair and stress-induced increases in plasma corticosterone levels

G protein-coupled receptors mediate transmission of information across the plasma membrane by activation of membrane-associated G-proteins that couple to intracellular effector systems. Although most G protein-coupled receptor ligands are confined to the extracellular space, for some G proteincoupled receptors, such as the thrombin receptor, thyrotropinreleasing hormone receptor, and the ␣ 2C -adrenoreceptor, a significant proportion of the receptor population is found in intracellular compartments. In the case of thrombin receptors, evidence suggests that the intracellular pool of receptors may serve as a reservoir, capable of restoring functional, noncleaved thrombin receptors to the plasma membrane after the cell has been exposed to thrombin (3). A minor fraction of thyrotropin-releasing hormone receptors expressed in HEK cells are targeted to the plasma membrane while a larger fraction is found in an intracellular compartment and are nonfunctional (4). However, when the thyrotropin releasing hormone receptors were expressed in two pituitary cell lines (GH3 and GHY cells) the receptors were found predominantly in the plasma membrane and to be functional by binding assays.For the ␣ 2C -adrenoreceptor, the functional status and role of an intracellular pool is less clear. This is particularly interesting in light of ␣ 2C

supporting

confidence: 48%

Section: Discussionmentioning

confidence: 99%

Cell-type Specific Targeting of the α2c-Adrenoceptor

Hurt¹,

Feng

Kobilka

2000

“…In these studies, ␣ 2C AR knockout mice performed less accurately in a delayed alternation task and displayed slowed motor initiation in the return phase of the task, supporting a role for the ␣ 2C AR in the cognitive aspect of response preparation. ␣ 2C AR knockout mice were also impaired in spatial and nonspatial water maze tests, thus supporting a role for this receptor in modulating cognitive functions (5), and alteration of ␣ 2C AR expression in transgenic mice has also been linked with behavioral despair development and changes in plasma corticosterone levels (11). Recent studies measuring [ 3 H]norepinephrine release from central neurons and cardiac sympathetic nerves have shown that the frequency-release curves for ␣ 2C AR-deficient mice are rightward shifted compared with wild-type mice (9).…”

Section: Resultsmentioning

confidence: 99%

A Four Amino Acid Deletion Polymorphism in the Third Intracellular Loop of the Human α2C-Adrenergic Receptor Confers Impaired Coupling to Multiple Effectors

Small

Forbes

Rahman

et al. 2000

177

166

The ␣ 2 -adrenergic receptors (␣ 2 ARs) play a critical role in modulating neurotransmitter release in the central and peripheral sympathetic nervous systems. A polymorphism of the ␣ 2 AR subtype localized to human chromosome 4 (the pharmacologic ␣ 2C AR subtype) within an intracellular domain has been identified in normal individuals. The polymorphism (denoted Del322-325) is because of an in-frame 12-nucleic acid deletion encoding a receptor lacking Gly-Ala-Gly-Pro in the third intracellular loop. To delineate the functional consequences of this structural alteration, Chinese hamster ovary cells were permanently transfected with constructs encoding wild-type human ␣ 2C AR and the polymorphic receptor. The Del322-325 variant had decreased high affinity agonist binding (K H ‫؍‬ 7.3 ؎ 0.95 versus 3.7 ؎ 0.43 nM; %R H ‫؍‬ 31 ؎ 4 versus 49 ؎ 4) compared with wild-type indicating impaired formation of the agonist-receptor-G protein complex. The polymorphic receptor displayed markedly depressed epinephrine-promoted coupling to G i , inhibiting adenylyl cyclase by 10 ؎ 4.3% compared with 73 ؎ 2.4% for wild-type ␣ 2C AR. This also was so for the endogenous ligand norepinephrine and full and partial synthetic agonists. Depressed agonist-promoted coupling to the stimulation of MAP kinase (ϳ71% impaired) and inositol phosphate production (ϳ60% impaired) was also found with the polymorphic receptor. The Del322-325 receptor was ϳ10 times more frequent in African-Americans compared with Caucasians (allele frequencies 0.381 versus 0.040). Given this significant loss of function phenotype in several signal transduction cascades and the skewed ethnic prevalence, Del322-325 represents a pharmacoethnogenetic locus and may also be the basis for interindividual variation in cardiovascular or central nervous system pathophysiology.The ␣ 2 -adrenergic receptors (␣ 2 AR) 1 are cell surface receptors for catecholamines, which couple to the G i /G o family of G proteins. ␣ 2 AR are expressed at multiple sites within the central and peripheral sympathetic nervous systems and may also be expressed at noninnervated sites of peripheral tissues as well. In the central nervous system presynaptic ␣ 2 AR act to inhibit the release of neurotransmitters such as norepinephrine, serotonin, and dopamine. As such, a number of responses have been ascribed to activation of these receptors by endogenous catecholamines or exogenously administered agonists. These include modulation of blood pressure, sedation, analgesia, opiate withdrawal, and multiple complex cognitive and behavioral parameters (1-5).Three human ␣ 2 AR subtypes have been cloned and characterized and are denoted as the ␣ 2A , ␣ 2B , and ␣ 2C subtypes (6 -8). Based on chromosomal localization, these have previously been denoted as ␣ 2 C10, ␣ 2 C2, and ␣ 2 C4, respectively. Recent studies including those with genetically engineered mice have shown that the ␣ 2C subtype plays specific roles in modulation of the acoustic startle reflex, prepulse inhibition, isolation-induced aggression, spatial...

“…␣ 2 AR are widely expressed within the central and peripheral nervous system (2-4) and participate in a broad spectrum of physiologic functions such as regulation of blood pressure both centrally and within the vasculature, sedation, analgesia, regulation of insulin release, renal function, and cognitive and behavioral functions (5)(6)(7)(8)(9)(10)(11)(12). Three human ␣ 2 AR subtypes have been cloned and characterized (␣ 2A , ␣ 2B , and ␣ 2C ).…”

mentioning

confidence: 99%

Polymorphic Deletion of Three Intracellular Acidic Residues of the α2B-Adrenergic Receptor Decreases G Protein-coupled Receptor Kinase-mediated Phosphorylation and Desensitization

Small

Brown

Forbes

et al. 2001

148

125

A polymorphic variant of the human ␣ 2B -adrenergic receptor (␣ 2B AR), which consists of a deletion of three glutamic acids (residues 301-303) in the third intracellular loop was found to be common in Caucasians (31%) and to a lesser extent in African-Americans (12%). The consequences of this deletion were assessed by expressing wild-type and the Del301-303 receptors in Chinese hamster ovary and COS cells. Ligand binding was not affected, although a small decrease in coupling efficiency to the inhibition of adenylyl cyclase was observed with the mutant. The deletion occurs within a stretch of acidic residues that is thought to establish the milieu for agonist-promoted phosphorylation and desensitization of the receptor by G protein-coupled receptor kinases (GRKs). Agonist-promoted phosphorylation studies carried out in cells coexpressing the ␣ 2B ARs and GRK2 revealed that the Del301-303 receptor displayed ϳ56% of wild-type phosphorylation. Furthermore, the depressed phosphorylation imposed by the deletion was found to result in a complete loss of short term agonistpromoted receptor desensitization. Thus the major phenotype of the Del301-303 ␣ 2B AR is one of impaired phosphorylation and desensitization by GRKs, and thus the polymorphisms renders the receptor incapable of modulation by this key mechanism of dynamic regulation.␣ 2 -Adrenergic receptors (␣ 2 AR) 1 are cell surface receptors for catecholamines that bind to the G i /G o family of G proteins, coupling to multiple effector systems including inhibition of adenylyl cyclase activity (1). ␣ 2 AR are widely expressed within the central and peripheral nervous system (2-4) and participate in a broad spectrum of physiologic functions such as regulation of blood pressure both centrally and within the vasculature, sedation, analgesia, regulation of insulin release, renal function, and cognitive and behavioral functions (5-12). Three human ␣ 2 AR subtypes have been cloned and characterized (␣ 2A , ␣ 2B , and ␣ 2C ). The ␣ 2B AR has a distinct pattern of expression within the brain, liver, lung, and kidney, and recent studies using gene knockouts in mice have shown that disruption of this receptor effects mouse viability (13), blood pressure responses to ␣ 2 AR agonists (13), and the hypertensive response to salt loading (14).Like the ␣ 2A AR subtype (15, 16), the ␣ 2B AR undergoes short term agonist promoted desensitization (17). This desensitization is due to phosphorylation of the receptor, which evokes a partial uncoupling of the receptor from functional interaction with G i /G o (18,19). Such phosphorylation appears to be due to G protein-coupled receptor kinases (GRKs), a family of serine/ threonine kinases that phosphorylate the agonist-occupied conformations of many G protein-coupled receptors (20). The process serves to finely regulate receptor function providing for rapid adaptation of the cell to its environment. Desensitization may also limit the therapeutic effectiveness of administered agonists. For the ␣ 2B AR, phosphorylation of serines/threonine...