Dopamine and new dopamine analogs: receptors and clinical applications

Division of dopamine (DA) receptors and alpha- and beta-adrenoceptors into two subtypes provides a pharmacological basis for the clinical use of DA and new DA receptor agonists in anesthesia and critical care medicine. First, differential receptor activation explains why three distinct cardiovascular and renal responses can be obtained at low, medium, and high infusion rates of DA. Low infusion rates, in which DA1 and DA2 receptors are activated, are being increasingly used to improve renal perfusion and to treat oliguric states. The medium dose range (activation of beta1-adrenoceptors) is used for treatment of heart failure. The high dose range (activation of alpha-adrenoceptors) is used for treatment of shock. Second, selective DA1 and relatively selective DA2 agonists and agonists with different combinations of DA and receptor activity other than DA have been synthesized and are being investigated for the treatment of congestive heart failure and hypertension. Some of these compounds could have advantages over DA for acute therapy. Future availability of these drugs in anesthesia and critical care settings will depend to a great extent on input from anesthesiologists concerning potential new uses and willingness to conduct clinical investigations.     

Dopamine, kidney, and hypertension: studies in dopamine receptor knockout mice

Dopamine is important in the pathogenesis of hypertension because of abnormalities in receptor-mediated regulation of renal sodium transport. Dopamine receptors are classified into D₁-like (D₁, D₅) and D₂-like (D₂, D₃, D₄) subtypes, all of which are expressed in the kidney. Mice deficient in specific dopamine receptors have been generated to provide holistic assessment on the varying physiological roles of each receptor subtype. This review examines recent studies on these mutant mouse models and evaluates the impact of individual dopamine receptor subtypes on blood pressure regulation.     

D1 dopamine receptor signaling involves caveolin-2 in HEK-293 cells

BACKGROUND: Dopamine receptors in the kidney, especially those belonging to the D1-like receptor family, are important in the regulation of renal function and blood pressure. Because of increasing evidence that G protein-coupled receptors (GPCRs) are associated with caveolae and lipid rafts, we tested the hypothesis that the D1 dopamine receptor (D1R) and signaling molecules are regulated by caveolin in caveolae or lipid rafts. METHODS: Six experimental approaches were used: (1) construction of tagged human D1Rs (hD1Rs) and transfectants; (2) cell culture [human embryonic kidney (HEK)-293 and immortalized rat renal proximal tubule cells] and biotinylation; (3) cell fractionation by sucrose gradient centrifugation; (4) immunoprecipitation and immunoblotting; (5) immunofluorescence and confocal microscopy; and (6) adenylyl cyclase assays. RESULTS: hD1Rs, heterologously expressed in HEK-293 cells, formed protein species with molecular mass ranging from 50 to 250 kD, and were localized in lipid rafts and nonraft plasma membranes. The hD1Rs cofractionated with caveolin-2, G protein subunits, and several signaling molecules. Both exogenously expressed hD1Rs and endogenously expressed rat D1Rs colocalized and coimmunoprecipitated with caveolin-2. A D1R agonist (fenoldopam) increased the amount of caveolin-2beta associated with hD1Rs and activated adenylyl cyclase to a greater extent in lipid rafts than in nonraft plasma membranes. Reduction in the expression of caveolin-2 with antisense oligonucleotides attenuated the stimulatory effect of fenoldopam on cyclic adenosine monophosphate (cAMP) accumulation. CONCLUSION: The majority of hD1Rs are distributed in lipid rafts. Heterologously and endogenously expressed D1Rs in renal cells are associated with and regulated by caveolin-2.     

Dopamine receptor subtypes in renal brush border and basolateral membranes

Dopamine (DA) modulates renal tubular sodium transport by actions at both brush border (BBM) and basolateral membranes (BLM). DA receptors have been demonstrated in proximal tubule but the subtype of DA receptor in either BBM or BLM has not been determined. DA-1 receptors were quantitated by the specific binding of 125I-SCH 23982, a DA-1 antagonist (defined by 20 microM SCH 23390, a DA-1 antagonist) and DA-2 receptors by the specific binding of 3H-methyl-spiroperidol or 3H-spiroperidol (defined by 30 microM trifluperidol, a predominantly DA-2 antagonist). The specific binding of 125I-SCH 23982 and 3H-methyl-spiroperidol or 3H-spiroperidol were saturable with time and ligand concentration and reversible. Analysis of Rosenthal plots by non-linear regression revealed a high affinity site and a very low affinity site for both BLM and BBM. Maximum receptor density was similar in BBM and BLM. Competition experiments with 125I-SCH 23982 revealed high and low affinity binding sites in both BBM and BLM. The high affinity site was characteristic of a DA-1 receptor. Competition experiments with 3H-spiroperidol were also suggestive of DA-2 receptors. DA-1 but not DA-2 drugs increased adenylate cyclase and phospholipase-C activities in both BBM and BLM. However, their effects were greater in BLM than BBM. We conclude that DA-1 and DA-2 receptors are present in both BBM and BLM in canine kidney. Renal DA-1 receptors are linked to stimulation of both adenylate cyclase and phospholipase-C activity.     

Dopaminergic defect in hypertension

Reverse genetics and the candidate gene approach have been utilized to identify the genetic defect(s) in hypertension. We have proposed the dopamine receptor gene as one candidate in the pathogenesis of hypertension. Because some forms of hypertension are sodium dependent or aggravated by sodium loading and because dopamine is important in aiding the organism to eliminate excess sodium, an abnormality in the renal dopaminergic system may be responsible for the sodium retention in hypertension. Both human and animal models of hypertension are associated with renal dopamine production and/or post first messenger defects. The Dahl salt-sensitive rat, which has a decreased ability to generate renal dopamine, and the spontaneously hypertensive rat (SHR), which has no such limitation, have a defective coupling of a D₁ receptor to a G protein/adenylyl cyclase complex. This coupling defect is: (1) genetic, since it precedes the onset of hypertension and co-segregates with the hypertensive phenotype, (2) receptor specific, since it is not shared by other humoral agents, and (3) organ and nephron segment selective, since it occurs in proximal tubules but not in cortical collecting ducts or the brain striatum. A consequence of the defective dopamine receptor/adenylyl cyclase coupling in the SHR is a decreased ability of D₁ agonists to inhibit Na⁺/H⁺ exchange activity. A resistance to the natriuretic effect of dopamine and D₁ agonists in the SHR is due mainly to decreased cyclic AMP production, although with maturation a post cyclic AMP defect is acquired. Radioligand binding studies suggest a loss of the high-affinity D₁ binding site in the SHR. However, sequencing of a limited segment of the D₁ receptor genes expressed in renal proximal tubule has not shown any difference between and the SHR and the normotensive Wistar Kyoto strain. Whether the defect is in the primary or tertiary structure of the receptor or due to a novel dopamine receptor that has not yet been cloned remains to be demonstrated.     

多巴胺D1、D2受体与觉醒及全身麻醉意识恢复的研究进展

背景 脑内多巴胺(dopamine,DA)及其受体参与觉醒、运动、认知和奖赏等行为调控,目前研究发现D1、D2受体也参与了全身麻醉意识恢复过程,但其具体机制尚未明确. 目的 探讨DA参与全身麻醉动物意识恢复的机制及其与D1、D2受体的关系. 内容 对DA D1和D2受体的结构、信号转导途径及其在中枢神经系统内的分布作一总结,对D1和D2受体参与全身麻醉后意识恢复中的机制研究进展作一综述. 趋向 D1、D2受体参与了全身麻醉动物意识恢复的过程,为研究和解释全身麻醉意识恢复机制提供新思路.     

Mitogen-activated protein kinase upregulation reduces renal D1 receptor affinity and G-protein coupling in obese rats

Reactive oxygen species play a key role in pathophysiology of cardiovascular diseases by modulating G-protein-coupled receptor signaling. We have shown that treatment of animal models of diabetes and aging with tempol decreases oxidative stress and restores renal dopamine D1 receptor (D1R) function. In present study, we determined whether oxidation of D1R and upregulation of mitogen-activated protein kinases (MAPK) were responsible for decreased D1R signaling in obese animals. Male lean and obese Zucker rats were supplemented with antioxidants tempol or lipoic acid for 2 weeks. Compared to lean, obese animals were hyperglycemic and hyperinsulinemic with increased oxidative stress, D1R oxidation and decreased glutathione levels. These animals had decreased renal D1R affinity and basal coupling to G-proteins. SKF-38393, a D1R agonist failed to stimulate G-proteins and adenylyl cyclase. Obese animals showed marked increase in renal MAPK activities. Treatment of obese rats with tempol or lipoic acid decreased blood glucose, reduced oxidative stress, and restored the basal D1R G-protein coupling. Antioxidants also normalized MAPK activities and restored D1R affinity and SKF-38393 induced D1R G-protein coupling and adenylyl cyclase stimulation. These studies show that D1R oxidation and MAPK upregulation contribute to D1R dysfunction in obese animals. Consequently, antioxidants while reducing the oxidative stress normalize the MAPK activities and restore D1R signaling.     

Dopamine "renal dose" versus fenoldopam mesylate to prevent ischemia-reperfusion injury in renal transplantation

Low dose of dopamine is commonly used after kidney transplantation as a reno-protective agent, although its benefits are controversial. Dopamine may increase renal blood flow, decrease resistive index (RI), and induce urine output in normal kidneys. Many authors hypothesized that the vasculature of a denervated renal transplant may not respond to dopamine in the same fashion as healthy native kidneys, which led us to find other drugs to attenuate the ischemia-reperfusion (I/R) injury. Fenoldopam is a selective dopamine1 (DA1) receptor agonist, most of the activity of which resides in the R-enantiomer, which also shows weaker alpha 2-adrenoceptor antagonist activities. Fenoldopam produces a vasidilatory effect in vascular beds that are rich in vascular DA1 receptors, producing increased renal blood flow at doses that do not affect blood pressure. In addition to its renal vasodilator activity, fenoldopam is natriuretic, possibly resulting from a direct effect of DA1 receptors on the proximal convoluted tubule. In animals with spontaneous or drug-induced renal failure, fenoldopam improves renal function. The aim of this study was to investigate the possible effects of fenoldopan mesylate in recent kidney transplants. Creatinine, blood urea nitrogen, urine output, and renal vascular resistive index (IR) were measured using Doppler ultrasound. Two groups of patients with no statistical differences in demographic data were treated with dopamine or fenoldopan, showing no significant difference but a trend favoring the fenoldopan group.     

Dopamine D(3) receptors and salt-dependent hypertension.

Alterations in the dopaminergic system may contribute to the pathogenesis of hypertension. Dopamine D(3) receptors have been shown to be involved in the regulation of sodium balance and hemodynamics in rodents. For determining the role of D(3) receptors in salt-dependent hypertension, clearance experiments were performed in anesthetized salt-sensitive (DS) and salt-resistant (DR) Dahl rats that were fed a standard diet with either normal (0.2%) or high (4%) sodium content for 21 to 26 d, which induced hypertension in DS but not in DR rats. The D(3) receptor agonist R(+)-7-hydroxydipropyl-aminotetralin (7-OH-DPAT) increased GFR by up to 35% and urinary sodium excretion by up to 4.4-fold in DR rats that were on both normal and high-sodium diet. 7-OH-DPAT-induced natriuresis also was observed in DS rats that were on normal diet but not in hypertensive DS rats that were on high-salt diet. No GFR response to 7-OH-DPAT was found in DS rats, irrespective of sodium diet. The diminished functional response to D(3) receptor stimulation in DS rats was associated with a significantly lower [(3)H]-7-OH-DPAT binding to renal membrane protein when comparing DS with DR rats. Consequently, DR rats were treated with BSF 135170, a novel, highly selective D(3) receptor antagonist, for 29 d. Whereas no change in systolic BP was observed during normal diet, high sodium intake significantly increased BP by almost 40 mmHg. In summary, both expression and function of the renal dopamine D(3) receptor are impaired in salt-sensitive Dahl rats. Together with the induction of salt-dependent hypertension in genetically salt-resistant Dahl rats by D(3) receptor blockade, the data strongly suggest that the deficiency in dopamine D(3) receptors represents an important pathophysiological factor in the development of salt-dependent hypertension.     

Characteristics of rat kidney dopamine receptors and the effects of renal denervation and dopamine infusion on these receptors

The characteristics of dopamine receptors, as well as the effects of denervation and dopamine infusion on dopamine receptors were studied using the radiolabeled receptor assay of [3H]-spiperone on rat kidney membrane preparations. The rat renal cortex was found to have a single class of [3H]-spiperone binding sites with a dissociation constant (Kd) of 13.5 +/- 2.2 nM. However, neither sulpiride nor serotonin strongly interacted with [3H]-spiperone binding, suggesting that DA1 receptors were predominant in the rat renal cortex. Denervation was performed by surgically stripping the nerves from the renal artery and coating them with 10% phenol. Chronic denervation had no significant effect on the affinity or maximum binding capacity of the renal dopamine receptors, although diuresis and the disappearance of catecholamine fluorophores in the denervated rats were observed. Chronic infusion of dopamine was performed using an osmotic minipump, resulting in a decrease in the number of rat kidney dopamine receptors. These results suggest that the dopamine receptor subtype in the renal cortex was mainly DA1, and that the major source of dopamine which affected the dopamine receptors in the rat kidney was not the nerve ending, but rather the circulation.     

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.     

Effect of low-dose dopamine on kidney function and vasoactive hormones in pediatric patients with advanced renal failure

Dopamine (DA) was infused in a dose of 2 micrograms/kg/min in 12 children and adolescents with chronic renal failure to test the vasodilatory reserve capacity of the kidney. Mean basal GFR and ERPF were 17.8 and 93.1 ml/min/1.73 m2, respectively. DA infusion had no significant influence on GFR but effective renal plasma flow (ERPF) increased by 14% (p less than 0.05). After DA, GFR did not correlate with ERPF. There was a significant increase in urinary sodium excretion (+22%). Sodium excretion correlated with osmotic clearance and urine flow rate with free water clearance. Plasma prolactin concentration was decreased (p less than 0.01), whereas noradrenaline, adrenaline and free dopamine increased significantly after DA. Plasma renin activity, aldosterone, arginine vasopressin and atrial natriuretic peptide levels remained unchanged. The data indicate that in pediatric patients with advanced renal failure DA fails to increase filtration capacity, whereas effective renal plasma flow and sodium excretion are stimulated. It is speculated that in this situation preglomerular and tubular renal functions regulated by dopamine receptors are better conserved than those affecting glomerular microcirculation.     

Mechanisms of disease: the role of GRK4 in the etiology of essential hypertension and salt sensitivity

Hypertension and salt sensitivity of blood pressure are two conditions the etiologies of which are still elusive because of the complex influences of genes, environment, and behavior. Recent understanding of the molecular mechanisms that govern sodium homeostasis is shedding new light on how genes, their protein products, and interacting metabolic pathways contribute to disease. Sodium transport is increased in the proximal tubule and thick ascending limb of Henle of the kidney in human essential hypertension. This Review focuses on the counter-regulation between the dopaminergic and renin-angiotensin systems in the renal proximal tubule, which is the site of about 70% of total renal sodium reabsorption. The inhibitory effect of dopamine is most evident under conditions of moderate sodium excess, whereas the stimulatory effect of angiotensin II is most evident under conditions of sodium deficit. Dopamine and angiotensin II exert their actions via G protein-coupled receptors, which are in turn regulated by G protein-coupled receptor kinases (GRKs). Polymorphisms that lead to aberrant action of GRKs cause a number of conditions, including hypertension and salt sensitivity. Polymorphisms in one particular member of this family-GRK4-have been shown to cause hyperphosphorylation, desensitization and internalization of a member of the dopamine receptor family, the dopamine 1 receptor, while increasing the expression of a key receptor of the renin-angiotensin system, the angiotensin II type 1 receptor. Novel diagnostic and therapeutic approaches for identifying at-risk subjects, followed by selective treatment of hypertension and salt sensitivity, might center on restoring normal receptor function through blocking the effects of GRK4 polymorphisms.     

Regulation of dopamine-induced natriuresisby the dopamine-metabolizing enzyme catechol-O-methyltransferase.

Dopamine (DA) is an intrarenal natriuretic hormone involved in sodium homeostasis. A study was performed to elucidate two possible regulatory pathways of DA-induced natriuresis, i.e., metabolism and precursor delivery. This was done by use of an intraperitoneal injection of a catechol-O-methyltransferase (COMT) inhibitor, entacapone, or intravenous infusion of the DA precursor, L-dopa. Entacapone (30 mg/kg i.p.) induced a more than fivefold increase in renal sodium excretion which occurred without changes in renal haemodynamics. The natriuretic response was highly dependent on DA D(1)-like receptor activation, since the selective D(1)-like receptor antagonist SCH23390 attenuated the natriuretic response by 61%, while the selective D(2)-like receptor antagonist sulpiride was ineffective. The urinary excretion of DA did not increase. Infusion of L-dopa (60 microg/h/kg) only induced a twofold increase in sodium excretion, but the urinary excretion of DA increased more than 17-fold. The L-dopa-induced natriuretic response occurred without increments in glomerular filtration rate and could be blocked with the D(1)-like receptor antagonist SCH23390. It is concluded that the DA-metabolizing enzyme COMT is involved in the regulation of the natriuretic effect of intrarenal DA. It may be speculated that intrarenal DA activity is not primarily determined on the basis of delivered precursor, but on that of the level of DA metabolism.     

Propofol decreases stimulated dopamine release in the rat nucleus accumbens by a mechanism independent of dopamine D2, GABAA and NMDA receptors

Although propofol (2,6-di-isopropylphenol) is a popular i.v. general anaesthetic, it has been suggested to have abuse potential. As many drugs of abuse act preferentially via release of dopamine in the limbic system, we investigated the action of propofol on stimulated dopamine release in the rat nucleus accumbens. Nucleus accumbens slices were superfused (1.6 ml min-1) with artificial cerebrospinal fluid at 32 degrees C. Dopamine release was evoked by electrical stimulation (10 pulses, 0.1 ms, 10 mA, 10 Hz, every 10 min) and monitored by fast cyclic voltammetry. Propofol 100 mumol litre-1 reduced stimulated dopamine release over the 2 h after administration, relative to intralipid controls, to mean 30 (SEM 2)% (P < 0.01). The dopamine D2 receptor antagonist metoclopramide 0.3 mumol litre-1 increased dopamine release but did not block the effect of propofol (38 (3)%). The selective GABAA antagonist bicuculline 24 mumol litre-1 also failed to antagonize the action of propofol (45 (3)%). The NMDA receptor antagonist dextromethorphan 10 mumol litre-1 decreased dopamine release to 57 (6)% (P < 0.01) but failed to block the inhibitory effect of propofol (46 (6)%). Although propofol has been reported to bind to D2, GABAA and NMDA receptors, failure of metoclopramide and bicuculline to block its effects suggests that an agonist action at D2 or GABAA receptors does not mediate the effects of propofol on dopamine release in the rat nucleus accumbens. The lack of effect of dextromethorphan makes an NMDA receptor antagonist action unlikely.      

Tempol reduces oxidative stress, improves insulin sensitivity, decreases renal dopamine D1 receptor hyperphosphorylation, and restores D1 receptor-G-protein coupling and function in obese Zucker rats

Oxidative stress plays a pathogenic role in hypertension, particularly the one associated with diabetes and obesity. Here, we test the hypothesis that renal dopamine D1 receptor dysfunction in obese Zucker rats is caused by oxidative stress. One group each from lean and obese Zucker rats received tempol, a superoxide dismutase mimetic in drinking water for 2 weeks. Obese animals were hypertensive, hyperglycemic, and hyperinsulinemic, exhibited renal oxidative stress, and increased protein kinase C activity. Also, there was hyperphosphorylation of D1 receptor, defective receptor-G-protein coupling, blunted dopamine-induced Na+-K+-ATPase inhibition, and diminished natriuretic response to D1 receptor agonist, SKF-38393. However, obese animals had elevated levels of plasma nitric oxide and urinary cGMP. In addition, L-N-nitroarginine and sodium nitroprusside showed similar effect on blood pressure in lean and obese rats. In obese animals, tempol reduced blood pressure, blood glucose, insulin, renal oxidative stress, and protein kinase C activity. Tempol also decreased D1 receptor phosphorylation and restored receptor G-protein coupling. Dopamine inhibited Na+-K+-ATPase activity, and SKF-38393 elicited a natriuretic response in tempol-treated obese rats. Thus in obese Zucker rats, tempol ameliorates oxidative stress and improves insulin sensitivity. Consequently, hyperphosphorylation of D1 receptor is reduced, leading to restoration of receptor-G-protein coupling and the natriuretic response to SKF-38393.     

Characterization of adenosine receptors in human kidney proximal tubule (HK-2) cells

Renal proximal tubule cells are particularly vulnerable to injury following ischemia and reperfusion due to their marginal blood supply and high metabolic demand. Renal adenosine receptor (AR) modulations preserve renal function following ischemic-reperfusion injury in vivo. Numerous intracellular proteins have been shown to be pivotal in the signal transduction of adenosine-mediated protection in vivo. However, characterization of the expression and function of ARs and intracellular proteins mediating protection in human proximal tubular cells is lacking. Therefore, we studied the ARs in an immortalized human renal proximal tubular cell (HK-2) line to determine if this cell line could function as an in vitro model of AR coupling. Immunoblotting with AR subtype specific antibodies detected all 4 subtypes of ARs (A(1), A(2a), A(2b) and A(3)), several isoforms of protein kinase C (alpha, delta, and epsilon and several heterotrimeric G-protein isoforms (G(i)alpha, G(s)alpha and G(q)alpha). The A(1) and A(3) ARs inhibited forskolin- stimulated adenylyl cyclase activity. The A(1) ARs also activated 42/44-kD ERK mitogen-activated protein kinases via G(i)- and tyrosine kinase-dependent pathways. The A(2a) ARs stimulated adenylyl cyclase activity and activated the protein kinase A-->CREB pathway. Chronic (48 h) treatment with a nonselective AR antagonist (8-phenyltheophylline) upregulated A(1), A(2a) ARs and G(i)alpha. Conversely, chronic stimulation of HK-2 ARs with a nonselective AR agonist (N-ethylcarbamoyladenosine) downregulated all 4 subtypes of ARs and G(s)alpha. Based on these findings, HK-2 cells are a useful in vitro model to study the signaling cascades of AR-mediated renal protection.