Common variants of the G protein-coupled receptor type 4 are associated with human essential hypertension and predict the blood pressure response to angiotensin receptor blockade.

Non-synonymous GRK4 variants, R65L, A142V and A486V, are associated with essential hypertension in diverse populations. This study replicated the association of GRK4 variants, including GRK4(142V), with human essential hypertension in a Japanese population (n=588; hypertensive, n=486 normotensive controls) and determined whether the presence of GRK4 variants predicted the blood pressure (BP) response to angiotensin receptor blockers (ARBs) in patients with essential hypertension. We analyzed 829 patients and compared the response to ARBs between individuals with no GRK4 variants (n=136) and those with variants at one or any of the three loci (n=693). Carriers of hGRK4(142V) had a greater decrease in systolic BP in response to ARBs than non-carrier hypertensive patients. By contrast, those with variants only at GRK4(486V) were less likely to achieve the BP goal in response to an ARB than those with no variants. These studies showed for the first time the association between GRK4(142V) and a larger decrease in BP with ARBs in hypertensive patients.

D1 dopamine receptor hyperphosphorylation in renal proximal tubules in hypertension.

A defect in the coupling of the D(1) receptor (D(1)R) to its G protein/effector complex in renal proximal tubules plays a role in the pathogenesis of spontaneous hypertension. As there is no mutation of the D(1)R gene in the spontaneously hypertensive rat (SHR), we tested the hypothesis that the coupling defect is associated with constitutive desensitization/phosphorylation of the D(1)R. The following experiments were performed: (1) Cell culture and membrane preparations from rat kidneys and immortalized rat renal proximal tubule cells (RPTCs); (2) immunoprecipitation and immunoblotting; (3) cyclic adenosine 3',5' monophosphate and adenylyl cyclase assays; (4) immunofluorescence and confocal microscopy; (5) biotinylation of cell surface proteins; and (6) in vitro enzyme dephosphorylation. Basal serine-phosphorylated D(1)Rs in renal proximal tubules, brush border membranes, and membranes from immortalized RPTCs were greater in SHRs (21.0+/-1.5 density units, DU) than in normotensive rats (7.4+/-2.9 DU). The increased basal serine phosphorylation of D(1)Rs in SHRs was accompanied by decreased expression of D(1)R at the cell surface, and decreased ability of a D(1)-like receptor agonist (fenoldopam) to stimulate cyclic adenosine 3',5' monophosphate (cAMP) production. Increasing protein phosphatase 2A activity with protamine enhanced the ability of fenoldopam to stimulate cAMP accumulation (17+/-4%) and alter D(1)R cell surface expression in intact cells from SHRs. Alkaline phosphatase treatment of RPTC membranes decreased D(1)R phosphorylation and enhanced fenoldopam stimulation of adenylyl cyclase activity (26+/-6%) in SHRs. Uncoupling of the D(1)R from its G protein/effector complex in renal proximal tubules in SHRs is caused, in part, by increased D(1)R serine phosphorylation.

Dopamine receptors in hypertension.

There is increased awareness of the role of dopamine in cardiovascular function, renal function and systemic blood pressure regulation. Growing evidence indicates that each of the five dopamine receptor subtypes participates in the regulation of blood pressure by mechanisms distinct for that particular subtype. Some dopamine receptors regulate blood pressure by influencing the central and peripheral nervous system, while others influence renal function and release of renin, aldosterone and vasopressin. This review summarizes the physiology and pathophysiology of the peripheral dopaminergic system and our current understanding of the role of individual dopamine receptors in the pathophysiology of human essential hypertension.

Adrenergic and endothelin B receptor-dependent hypertension in dopamine receptor type-2 knockout mice.

Polymorphism of the dopamine receptor type-2 (D(2)) gene is associated with essential hypertension. To assess whether D(2) receptors participate in regulation of blood pressure (BP), we studied mice in which the D(2) receptor was disrupted. In anesthetized mice, systolic and diastolic BPs (in millimeters of mercury) were higher in D(2) homozygous and heterozygous mutant mice than in D(2)+/+ littermates. BP after alpha-adrenergic blockade decreased to a greater extent in D(2)-/- mice than in D(2)+/+ mice. Epinephrine excretion was greater in D(2)-/- mice than in D(2)+/+ mice, and acute adrenalectomy decreased BP to a similar level in D(2)-/- and D(2)+/+ mice. An endothelin B (ET[B]) receptor blocker for both ET(B1) and ET(B2) receptors decreased, whereas a selective ET(B1) blocker increased, BP in D(2)-/- mice but not D(2)+/+ mice. ET(B) receptor expression was greater in D(2)-/- mice than in D(2)+/+ mice. In contrast, blockade of ET(A) and V(1) vasopressin receptors had no effect on BP in either D(2)-/- or D(2)+/+ mice. The hypotensive effect of an AT(1) antagonist was also similar in D(2)-/- and D(2)+/+ mice. Basal Na(+),K(+)-ATPase activities in renal cortex and medulla were higher in D(2)+/+ mice than in D(2)-/- mice. Urine flow and sodium excretion were higher in D(2)-/- mice than in D(2)+/+ mice before and after acute saline loading. Thus, complete loss of the D(2) receptor results in hypertension that is not due to impairment of sodium excretion. Instead, enhanced vascular reactivity in the D(2) mutant mice may be caused by increased sympathetic and ET(B) receptor activities.

Impaired renal D(1)-like and D(2)-like dopamine receptor interaction in the spontaneously hypertensive rat.

D(1)-like (D(1), D(5)) and D(2)-like (D(2), D(3), D(4)) dopamine receptors interact in the kidney to produce a natriuresis and a diuresis. Disruption of D(1) or D(3) receptors in mice results in hypertension that is caused, in part, by a decreased ability to excrete an acute saline load. We studied D(1)-like and D(2)-like receptor interaction in anesthetized spontaneously hypertensive rats (SHR) by the intrarenal infusion of Z-1046 (a novel dopamine receptor agonist with rank order potency of D(3)> or =D(4)>D(2)>D(5)>D(1)). Z-1046 increased glomerular filtration rate (GFR), urine flow, and sodium excretion in normotensive Wistar-Kyoto rats but not in SHRs. The lack of responsiveness to Z-1046 in SHRs was not an epiphenomenon, because intrarenal cholecystokinin infusion increased GFR, urine flow, and sodium excretion to a similar extent in the two rat strains. We conclude that renal D(1)-like and D(2)-like receptor interaction is impaired in SHRs. The impaired D(1)-like and D(2)-like receptor interaction in SHRs is not caused by alterations in the coding sequence of the D(3) receptor, the D(2)-like receptor expressed in rat renal tubules that has been shown to be involved in sodium transport. Because the diuretic and natriuretic effects of D(1)-like receptors are, in part, caused by an interaction with D(2)-like receptors, it is possible that the decreased Z-1046 action in SHRs is secondary to the renal D(1)-like receptor dysfunction in this rat strain.

D(1) dopamine receptor regulation of NHE3 during development in spontaneously hypertensive rats.

To determine if the defective interactions among D(1)-like receptors, G proteins, and Na(+)/H(+) exchanger 3 (NHE3) are consequences of hypertension, we studied these interactions in rats, before (2--3 wk) and after (12 wk) the establishment of hypertension. To eliminate the confounding influence of second messenger action on D(1) receptor-NHE3 interaction, studies were performed in renal brush-border membranes (BBM) devoid of cytoplasmic second messengers. NHE3 activity increased with age in Wistar-Kyoto (WKY) rats (3 wk = 1.48 +/- 0.39, n = 13; 12 wk = 2.83 +/- 0.15, n = 16, P < 0.05) but not in spontaneously hypertensive rats (SHRs; 3 wk = 2.52 +/- 0.37, n = 11; 12 wk = 2.81 +/- 0.20, n = 16). D(1) receptor protein tended to decrease, whereas NHE3 protein tended to increase with age in both WKY and SHRs. However, the inhibitory effect of a D(1)-like agonist, SKF-81297, on NHE3 activity increased with age in WKY rats (3 wk = -40.7 +/- 5.3%, n = 10, 12 wk = -58.7 +/- 4.6%, n = 12, P < 0.05) but not in SHRs (3 wk = -27.6 +/- 5.9%, n = 11, 12 wk = -25.1 +/- 3.2%, n = 11). The decreased inhibitory effect of another D(1)-like agonist, fenoldopam, on NHE3 activity in SHRs was not caused by increased activity and binding of G beta gamma to NHE3 as has been reported in young WKY rats. G(s)alpha mediates, in part, the inhibitory effect of D(1)-like agonists on NHE3 activity. In WKY rats, fenoldopam increased G(s)alpha/NHE3 binding to the same extent in 2-wk-old (1.5-fold, n = 4) and adult (1.5-fold, n = 4) rats. In contrast, in SHRs, fenoldopam decreased the amount of G(s)alpha bound to NHE3 in 2-wk-old SHRs and had no effect in 4-wk-old and adult SHRs. These studies indicate that the decreased inhibitory effect of D(1)-like agonists on NHE3 activity in SHRs (compared with WKY rats) precedes the development of hypertension. This may be caused, in part, by a decreased interaction between G(s)alpha and NHE3 in BBM secondary to impaired D(1)-like receptor function.

[Regulation of NHE3 by D1 dopamine receptor during development of spontaneously hypertensive rats].

OBJECTIVE: To determine if spontaneous hypertension is secondary to defective interaction among dopamine receptor, G protein, and Na(+)/H(+) exchanger 3 (NHE3). METHODS: The inhibitory effect of a D(1) dopamine agonist upon NHE3 activity and its impact upon G(s)alpha/NHE3 binding in renal brush border membrane (BBM) of spontaneously hypertensive rats (SHRs) 2 - 3 weeks before and 12 weeks after the establishment of hypertension were examined. In order to avoid the confounding influence of second messenger on D(1) receptor/NHE3 interaction, study was made in BBM devoid of cytoplasmic component. RESULTS: NHE3 activity increased with age in Wister-Kyoto (WKY) rats but not in SHRS. D1 receptor expression did change with age in both WKY rats and SHRs. The inhibitory effect of a D(1)-like agonist on NHE3 activity increased with age in WKY rats but not in SHRs. In WKY rats, another D(1)-like agonist, fedoldopam, increased G(s)alpha/NHE3 binding to the same extent in 2 week old and adult rats, but decreased the amount of G(s)(alpha)bound to NHE3 in 2 week old SHRs. CONCLUSION: The decrease of inhibitory effect of D(1)-like agonist upon NHE3 activity in SHRs precedes the development of hypertension. Spontaneous hypertension may be caused, in part, by a decreased interaction between G(s)alpha and NHE3 in BBM secondary to D(1)-like receptor function.

Renal dopamine and sodium homeostasis.

During the past decade, it has become evident that dopamine plays an important role in the regulation of fluid and electrolyte balance and blood pressure. Dopamine exerts its actions through two families of dopamine receptors, designated D1-like and D2-like, which are identical in the brain and in peripheral tissues. The two D1-like receptors--D1 and D5 receptors--expressed in mammals are linked to stimulation of adenylyl cyclase. The three D2-like receptors--D2, D3, and D4,--are linked to inhibition of adenylyl cyclase. Dopamine affects fluid and electrolyte balance by regulation of renal excretion of electrolytes and water through actions on renal hemodynamics and tubular epithelial transport and by modulation of the secretion and/or action of vasopressin, renin, aldosterone, catecholamines, and endothelin B receptors (ETB) receptors. It also affects fluid and sodium intake by way of "appetite" centers in the brain and alterations of gastrointestinal tract transport. The production of dopamine in neural and non-neural tissues and the presence of receptors in these tissues suggest that dopamine can act in an autocrine or paracrine fashion. This renal autocrine-paracrine function, which becomes most evident during extracellular fluid volume expansion, is lost in essential hypertension and in some animal models of genetic hypertension. This deficit may be caused by abnormalities in renal dopamine production and polymorphisms or abnormal post-translational modification and regulation of dopamine receptor subtypes.

Gbeta regulation of Na/H exchanger-3 activity in rat renal proximal tubules during development.

The decreased natriuretic action of dopamine in the young has been attributed to decreased generation of cAMP by the activated renal D(1)-like receptor. However, sodium/hydrogen exchanger (NHE) 3 activity in renal brush-border membrane vesicles (BBMV) can be modulated independent of cytoplasmic second messengers. We therefore studied D(1)-like receptor regulation of NHE activity in BBMVs in 2-, 4-, and 12-wk-old (adult) rats. Basal NHE activity was least in 2-wk-old compared with 4- and 12-wk-old rats. D(1)-like agonist (SKF-81297) inhibition of NHE activity was also least in 2-wk-old (-1 +/- 9%, n = 3) compared with 4 (-15 +/- 5%, n = 6)- and 12 (-65 +/- 4%, n = 6)-wk-old rats. The decreased response to the D(1)-like agonist in BBMV was not caused by decreased D(1) receptors or NHE3 expression in the young. G(s)alpha, which inhibits NHE3 activity by itself, coimmunoprecipitated with NHE3 to the same extent in 2-wk-old and adult rats. G(s)alpha function was also not impaired in the young because guanosine 5'-O-(3-thiotriphosphate) decreased NHE activity to a similar extent in 4-wk-old and adult rats. Galpha(i-3) protein expression in BBMV also did not change with age. In contrast, Gbeta expression and the amount of Gbeta that coimmunoprecipitated with NHE3 in BBMV was greatest in 2-wk-old rats and decreased with age. Gbeta common antibodies did not affect D(1)-like agonist inhibition of NHE activity in adult rats (8%) but markedly increased it (48%)in 4-wk-old rats. We conclude that the decreased inhibitory effect of D(1)-like receptors on NHE activity in BBMV in young rats is caused, in part, by the increased expression and activity of the G protein subunit Gbeta/gamma. The direct regulation of NHE activity by G protein subunits may be an important step in the maturation of renal tubular ion transport.

D1 dopamine receptor signalling defect in spontaneous hypertension.

Dopamine modulates cardiovascular function by actions in the central and peripheral nervous system, by altering the secretion/release of prolactin, pro-opiomelanocortin, vasopressin, aldosterone, and renin, and by directly affecting renal function. Dopamine produced by the renal proximal tubule exerts an autocrine/paracrine action via two classes of dopamine receptors, D1-like (D1 and D5) and D2-like (D2, D3, and D4), that are differentially expressed along the nephron. The autocrine/paracrine function of dopamine, manifested by tubular rather than by haemodynamic mechanisms, becomes most evident during extracellular fluid volume expansion. This renal autocrine/paracrine function is lost in essential hypertension and in some animal models of genetic hypertension. The molecular basis for the dopaminergic dysfunction in hypertension may involve an abnormal post-translational modification of dopamine receptors.

Renal protein phosphatase 2A activity and spontaneous hypertension in rats.

The impaired renal paracrine function of dopamine in spontaneously hypertensive rats (SHR) is caused by hyperphosphorylation and desensitization of the renal D(1) dopamine receptor. Protein phosphatase 2A (PP(2A)) is critical in the regulation of G-protein-coupled receptor function. To determine whether PP(2A) expression and activity in the kidney are differentially regulated in genetic hypertension, we examined the effects of a D(1)-like agonist, fenoldopam, in renal cortical tubules and immortalized renal proximal tubule cells from normotensive Wistar-Kyoto rats (WKY) and SHR. In cortical tubules and immortalized proximal tubule cells, PP(2A) expression and activities were greater in cytosol than in membrane fractions in both WKY and SHR. Although PP(2A) expressions were similar in WKY and SHR, basal PP(2A) activity was greater in immortalized proximal tubule cells of SHR than WKY. In immortalized proximal tubule cells of WKY, fenoldopam increased membrane PP(2A) activity and expression of the regulatory subunit PP(2A)-B56alpha, effects that were blocked by the D(1)-like antagonist SCH23390. Fenoldopam had no effect on cytosolic PP(2A) activity but decreased PP(2A)-B56alpha expression. In contrast, in immortalized proximal tubule cells of SHR, fenoldopam decreased PP(2A) activity in both membranes and cytosol but predominantly in the membrane fraction, without affecting PP(2A)-B56alpha expression; this effect was blocked by the D(1)-like antagonist SCH23390. We conclude that renal PP(2A) activity and expression are differentially regulated in WKY and SHR by D(1)-like receptors. A failure of D(1)-like agonists to increase PP(2A) activity in proximal tubule membranes may be a cause of the increased phosphorylation of the D(1) receptor in the SHR.

Role of dopamine in the pathogenesis of hypertension.

1. Dopamine, via different dopamine receptor subtypes, regulates cardiovascular functions by actions on the central and peripheral nervous systems, vascular smooth muscle, the heart and the kidney. The dopaminergic system in the central nervous system (CNS) may participate in the regulation of systemic blood pressure. 2. Dopamine 'D2-like' (D2, D3 and D4) receptors, rather than 'D1-like' (D1 and D5) receptors, are involved in the CNS regulation of blood pressure; post-synaptic D2-like receptors increase blood pressure, while presynaptic D2-like receptors (the predominant action) produce the opposite effect. 3. Outside the CNS, dopamine may regulate blood pressure via pressure controls that act with intermediate rapidity (e.g. stress relaxation, arginine vasopressin and renin-angiotensin vasoconstriction), as well as those systems related to the long-term control of body fluid volume. 4. Dopamine D1- and D2-like receptors have been described in resistance vessels, such as the renal, mesenteric, coronary, pulmonary and cerebral arteries. The ability of D1-like receptors to inhibit renal smooth muscle hypertrophy indicates their importance in longer-term regulation of blood pressure. 5. Aberrant dopaminergic regulation of aldosterone secretion, via D2-like receptors, has been reported to be involved in some forms of hyperaldosteronism and hypertension. Some forms of hypertension may also be caused by an aberrant renal dopaminergic system. Abnormalities of three aspects of the renal dopaminergic system may lead to hypertension: (i) renal production of dopamine; (ii) transduction of the renal vascular dopamine signal; and (iii) transduction of the renal tubular dopamine signal. 6. Thus, increased blood pressure occurs after either blockade of D1-like receptors or of dopamine production in rats or disruption of the D1 receptor or the D3 receptor gene in mice.

Renal dopamine receptors in health and hypertension.

During the past decade, it has become evident that dopamine plays an important role in the regulation of renal function and blood pressure. Dopamine exerts its actions via a class of cell-surface receptors coupled to G-proteins that belong to the rhodopsin family. Dopamine receptors have been classified into two families based on pharmacologic and molecular cloning studies. In mammals, two D1-like receptors that have been cloned, the D1 and D5 receptors (known as D1A and D1B, respectively, in rodents), are linked to stimulation of adenylyl cyclase. Three D2-like receptors that have been cloned (D2, D3, and D4) are linked to inhibition of adenylyl cyclase and Ca2+ channels and stimulation of K+ channels. All the mammalian dopamine receptors, initially cloned from the brain, have been found to be expressed outside the central nervous system, in such sites as the adrenal gland, blood vessels, carotid body, intestines, heart, parathyroid gland, and the kidney and urinary tract. Dopamine receptor subtypes are differentially expressed along the nephron, where they regulate renal hemodynamics and electrolyte and water transport, as well as renin secretion. The ability of renal proximal tubules to produce dopamine and the presence of receptors in these tubules suggest that dopamine can act in an autocrine or paracrine fashion; this action becomes most evident during extracellular fluid volume expansion. This renal autocrine/paracrine function is lost in essential hypertension and in some animal models of genetic hypertension; disruption of the D1 or D3 receptor produces hypertension in mice. In humans with essential hypertension, renal dopamine production in response to sodium loading is often impaired and may contribute to the hypertension. The molecular basis for the dopaminergic dysfunction in hypertension is not known, but may involve an abnormal post-translational modification of the dopamine receptor.

Effects of costimulation of dopamine D1- and D2-like receptors on renal function.

In vitro studies have suggested that dopamine D1- and D2-like receptors interact to inhibit renal sodium transport. We used Z-1046, a dopamine receptor agonist with the rank-order potency D3 >/= D4 > D2 > D5 > D1, to test the hypothesis that D1- and D2-like receptors interact to inhibit renal sodium transport in vivo in anesthetized rats. Increasing doses of Z-1046, administered via the right renal artery, increased renal blood flow (RBF), urine flow, and absolute and fractional sodium excretion without affecting glomerular filtration rate. For determination of the dopamine receptor involved in the renal functional effects of Z-1046, another group of rats received Z-1046 at 2 microgram . kg-1 . min-1 (n = 10) in the presence or absence of the D2-like receptor antagonist domperidone and/or the D1-like antagonist SCH-23390. Domperidone alone had no effect but blocked the Z-1046-mediated increase in urine flow and sodium excretion; it enhanced the increase in RBF after Z-1046. SCH-23390 by itself decreased urine flow and sodium excretion without affecting RBF and blocked the diuretic, natriuretic, and renal vasodilatory effect of Z-1046. We conclude that the renal vasodilatory effect of Z-1046 is D1-like receptor dependent, whereas the diuretic and natriuretic effects are both D1- and D2-like receptor dependent.

Disruption of the dopamine D3 receptor gene produces renin-dependent hypertension.

Since dopamine receptors are important in the regulation of renal and cardiovascular function, we studied the cardiovascular consequences of the disruption of the D3 receptor, a member of the family of D2-like receptors, expressed in renal proximal tubules and juxtaglomerular cells. Systolic and diastolic blood pressures were higher (approximately 20 mmHg) in heterozygous and homozygous than in wild-type mice. An acute saline load increased urine flow rate and sodium excretion to a similar extent in wild-type and heterozygous mice but the increase was attenuated in homozygous mice. Renal renin activity was much greater in homozygous than in wild-type mice; values for heterozygous mice were intermediate. Blockade of angiotensin II subtype-1 receptors decreased systolic blood pressure for a longer duration in mutant than in wild-type mice. Thus, disruption of the D3 receptor increases renal renin production and produces renal sodium retention and renin-dependent hypertension.

Renal nerves and D1-dopamine receptor-mediated natriuresis.

The resistance of the spontaneously hypertensive rat (SHR) kidney to the natriuretic effect of dopamine and D1 agonists may be due to increased renal nerve activity. Therefore, we compared the effects of the intrarenal arterial infusion of the D1 agonist, SKF 38383, into the denervated (DNX) kidney of saline-loaded-anesthetized SHR and its control, the Wistar-Kyoto (WKY) rat. In both WKY and SHR, DNX of the left kidney slightly decreased urine flow (UV) and absolute (UNaV) and fractional sodium excretion (FENa) in the innervated right kidney; neither vehicle nor D1 agonist infusion exerted any effect. In the left kidney, denervation increased UV, UNaV, and FENa to a similar degree in WKY and SHR (2-fold), without affecting renal blood flow, glomerular filtration rate, or blood pressure. In WKY but not in SHR, after DNX, the D1 agonist dose-dependently increased UV, UNaV, and FENa in the denervated kidney. We conclude that the decreased natriuretic effect of D1 agonists in the SHR is not due to increased renal nerve activity. These data support our previous studies implicating a defect of the D1 receptor or its regulation in the kidney in genetic hypertension.

Dopamine and diltiazem-induced natriuresis in the spontaneously hypertensive rat.

An attenuated natriuretic response to dopamine and D1 agonists in genetic hypertension has been attributed to an uncoupling of the renal D1 dopamine receptor from its G protein-effector protein complex. We have reported that in normotensive Wistar-Kyoto (WKY) rats the natriuresis induced by calcium channel blockers is caused in part by activation of renal D1 dopamine receptors. We tested the interaction between the renal D1 receptor and a calcium channel blocker, diltiazem, infused into a renal artery of anesthetized spontaneously hypertensive rats (SHR) acutely loaded with 5% saline. Diltiazem produced a 50% increase in renal blood flow and nearly tripled absolute and fractional sodium excretion; urine flow rate more than doubled, but glomerular filtration rate did not change. However, the D1 receptor antagonist SKF-83742, which had no effect by itself, did not diminish the response to diltiazem. In a separate group of concurrent experiments, we found that the diltiazem-induced natriuresis was associated with a decrease in Na(+)-K(+)-adenosinetriphosphatase activity in the renal medulla of SHR. In contrast, in WKY rats, no changes were noted in the renal medulla but a decrease in Na(+)-K(+)-adenosinetriphosphatase activity was noted in the renal cortex. Diltiazem had no effect on urinary dopamine excretion in either rat strain. We conclude that diltiazem induces natriuresis differently in SHR and WKY rats; it is independent of D1 receptors in SHR and is in great part mediated by renal hemodynamic, rather than by cortical tubular, effects. These studies support previous findings of a defective renal cortical tubular D1 mechanism in SHR.

Transgenic mice to study the role of dopamine receptors in cardiovascular function.

Dopamine, an intrarenal regulator of sodium transport, is important in the pathogenesis of hypertension. The transduction of D1-like receptors in renal proximal tubules is defective in animal models of genetic hypertension. The defect is associated with an impaired regulation of proximal tubular sodium transport and cosegregates with hypertension in rats. Moreover, mice lacking one or both D1A receptor alleles develop hypertension. Extrasynaptic D3 receptors in renal tubules and juxtaglomerular cells may also regulate renal sodium transport and renin secretion while presynaptic D3 receptors may act as autoreceptors to inhibit neural norepinephrine release. Mice lacking one or both D3 alleles have elevated systolic blood pressure and developed diastolic hypertension. Although basal urine flow, sodium excretion, and glomerular filtration rate are similar, mice homozygous to the D3 receptor have an impaired ability to excrete an acute saline load compared to heterozygous and wild type mice. These studies suggest that abnormalities in dopamine receptor genes or their regulation may lead to the development of hypertension via different pathogenetic mechanisms.

Dopamine D1A receptor regulation of phospholipase C isoform.

In LTK- cells stably transfected with rat D1A receptor cDNA, fenoldopam, a D1 agonist, increased phosphatidylinositol 4, 5-bisphosphate hydrolysis in a time-dependent manner. In the cytosol, phospholipase C (PLC) activity increased (50 +/- 7%) in 30 s, returned to basal level at 4 h, and decreased below basal values by 24 h; in the membrane, PLC activity also increased (36 +/- 13%) in 30 s, returned to basal level at 10 min, and decreased below basal value at 4 and 24 h. Fenoldopam also increased PLC-gamma protein in a time-dependent manner. The latter was blocked by the D1 antagonist SKF83742 and by a D1A antisense oligodeoxynucleotide, indicating involvement of the D1A receptor. The fenoldopam-induced increase in PLC-gamma and activity was mediated by protein kinase A (PKA) since it was blocked by the PKA antagonist Rp-8-CTP-adenosine cyclic 3':5'-monophosphorothioate (Rp-8-CTP-cAMP-S) and mimicked by direct stimulation of adenylyl cyclase with forskolin or by a PKA agonist, Sp-cAMP-S. Protein kinase C (PKC) was also involved, since the fenoldopam-induced increase in PLC-gamma protein was blocked by two different PKC inhibitors, calphostin C and chelerythrine; calphostin C also blocked the fenoldopam-induced increase in PLC activity. In addition, forskolin and a PKA agonist, Sp-8-CTP-cAMP-S, increased PKC activity, and direct stimulation of PKC with phorbol 12-myristate 13-acetate increased PLC-gamma protein and activity, effects that were blocked by calphostin C. We suggest that the D1A-mediated stimulation of PLC occurs as a result of PKA activation. PKA then stimulates PLC-gamma in cytosol and membrane via activation of PKC.