Renal Dopamine Receptors and Oxidative Stress: Role in Hypertension

Abstract

Significance: The kidney plays an important role in the long-term control of blood pressure. Oxidative stress is one of the fundamental mechanisms responsible for the development of hypertension. Dopamine, via five subtypes of receptors, plays an important role in the control of blood pressure by various mechanisms, including the inhibition of oxidative stress.

Recent Advances: Dopamine receptors exert their regulatory function to decrease the oxidative stress in the kidney and ultimately maintain normal sodium balance and blood pressure homeostasis. An aberration of this regulation may be involved in the pathogenesis of hypertension.

Critical Issues: Our present article reviews the important role of oxidative stress and intrarenal dopaminergic system in the regulation of blood pressure, summarizes the current knowledge on renal dopamine receptor-mediated antioxidation, including decreasing reactive oxygen species production, inhibiting pro-oxidant enzyme nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase, and stimulating antioxidative enzymes, and also discusses its underlying mechanisms, including the increased activity of G protein-coupled receptor kinase 4 (GRK4) and abnormal trafficking of renal dopamine receptors in hypertensive status.

Future Directions: Identifying the mechanisms of renal dopamine receptors in the regulation of oxidative stress and their contribution to the pathogenesis of hypertension remains an important research focus. Increased understanding of the role of reciprocal regulation between renal dopamine receptors and oxidative stress in the regulation of blood pressure may give us novel insights into the pathogenesis of hypertension and provide a new treatment strategy for hypertension.

Introduction

It is well accepted that hypertension is a complex trait determined by genetic, epigenetic, behavioral, and environmental factors and their intricate interactions. The kidney plays an important role in the long-term control of blood pressure and is the major organ involved in the regulation of sodium homeostasis (60). An inappropriate sodium retention in hypertension results from enhanced renal sodium transport per se or a failure to respond appropriately to signals that decrease renal sodium transport in the face of increased sodium intake. Sodium excretion is regulated by numerous humoral or hormone factors. Among these, dopamine has been shown to be an important regulator of sodium balance and blood pressure through an independent renal dopaminergic system (187a, 195). Oxidative stress has gained attention as one of the fundamental mechanisms involved in the development of hypertension (42, 101). Our studies and those of others have shown that the function of renal dopamine receptors is impaired in hypertension, in part, because of oxidative stress (96, 116, 165). In this article, we review the role of oxidative stress and intrarenal dopaminergic system in the regulation of blood pressure, the abnormality of this interaction in hypertension, and the current knowledge of the mechanisms involved in this interaction. These may advance our understanding of the role of reciprocal regulation between renal dopamine receptors and oxidative stress in the control of blood pressure and highlight potential and novel strategies for the prevention and treatment of hypertension.

Reactive Oxygen Species and Hypertension

The term oxidative stress can be defined as a serious disturbance of the balance between the production of reactive oxygen species (ROS) and the antioxidant defense systems, and has been shown in a wide range of studies to contribute to the pathogenesis of many diseases, such as hypertension (34). ROS are a class of oxygen atoms that contain unpaired electrons, including superoxide anion (O₂⁻), hydrogen peroxide (H₂O₂), hydroxyl anion (⁻OH) formed from H₂O₂, and singlet oxygen (¹O₂), hypochlorous acid (HClO), among others, together with secondary products of their interaction with nitric oxide (NO) and lipids. ROS are naturally generated by specific oxidases, such as the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase, xanthine/xanthine oxidase, various arachidonic acid monooxygenases, and enzymes of the mitochondrial respiratory chain (50, 85) (Fig. 1). Apart from the above, numerous other enzymes such as NO synthase, cyclooxygenases, cytochrome P450 enzymes, and lipoxygenases, as well as some cell organelles such as the peroxisome and endoplasmic reticulum, also contribute to intracellular ROS production (37, 66). Moreover, there is cross talk among these pro-oxidant systems, which leads to increased levels of ROS (38). Mitochondrial respiration is a major site of ROS generation, whereas membrane-associated NADPH oxidase is the most important source of O₂⁻ outside the mitochondria in nonphagocytic cells, including vascular and renal mesangial and tubular cells (39, 127). However, the levels of ROS depend not only on their production but also on their degradation by antioxidant enzymes. While superoxide dismutase (SOD) is one of the major defense systems to remove ROS, catalase (CAT), peroxiredoxins (Prxs), glutathione peroxidase (GSH-Px), glutathione transferase (GST), glutathione reductase (GR), and thioredoxin (Trx) reductase are all important in the metabolism of H₂O₂ (72, 112). In addition, some nonenzymatic compounds such as ascorbate, β-carotene, glutathione (GSH), nicotinamide, and tocopherol/vitamin E also counteract ROS (129). Once oxidation exceeds antioxidation, the balance between the oxidation system and antioxidant system is broken, oxidative stress occurs, causing deleterious effects, including tissue damage.

FIG. 1.

Schematic diagram of ROS production and metabolism. Multiple sources can produce superoxide (O₂^•−), which can rapidly react with NO to form peroxynitrite (ONOO^•) or be converted to H₂O₂ via SOD. H₂O₂ can be converted to water (H₂O) and O₂ via catalase, glutathione peroxidase, or thioredoxin peroxidase. H₂O₂ can also produce hydroxyl radical (OH^•) through the Fenton reaction, or generate hypochlorous acid (HClO) via the enzyme myeloperoxidase. AA, arachidonic acid; Cyt, cytochrome; H₂O₂, hydrogen peroxide; Mito, mitochondrial; NADPH, nicotinamide-adenine dinucleotide phosphate; NO, nitric oxide; ROS, reactive oxygen species; SOD, superoxide dismutase.

Role of oxidative stress in hypertension

In 1990, Schneider et al. first reported that hypertensive patients had lower glutathione peroxide activity and higher SOD and serum GR activity compared with normotensive healthy subjects (126). In 1991, oxidative stress was implicated to play an important role in the pathogenesis of hypertension. Nakazono et al. reported that the intravenous injection of a fusion protein consisting of human Cu/Zn-type SOD and a C-terminal basic peptide, targeted to endothelial cells, decreased the blood pressure of spontaneously hypertensive rats (SHRs), but not normotensive rats (104). Subsequently, a number of studies have shown the vital role of ROS in the development of hypertension (1, 148). Excessive ROS generation and/or impaired antioxidant mechanisms are commonly observed in various animal models of genetic and experimental hypertension (26, 49, 163). These were confirmed in specific ROS generating gene-knockout mice, or animal models of ROS-mediated hypertension, in which ROS production is suppressed. Mice models with genetic deficiency of superoxide-generating enzymes, such as NADPH oxidase NOX1 or NOX2, have decreased blood pressure (51, 128). Inhibition of ROS generation with apocynin, a NADPH oxidase inhibitor, or allopurinol, a xanthine oxidase inhibitor, and radical scavenging with antioxidants such as resveratrol or SOD mimetics decrease blood pressure and prevent the development of hypertension in rodent models of hypertension (31, 57, 118). Moreover, the association between ROS and increased blood pressure is also demonstrated in hypertensive human subjects. Clinical studies have shown increased ROS production in patients with various types of hypertension, including essential hypertension, renovascular hypertension, malignant hypertension, and preeclampsia (64, 142, 144). All of these reports contribute to the accumulating evidence that increased oxidative stress is involved in the pathogenesis of hypertension.

Studies have shown that oxidative stress induces hypertension through numerous mechanisms in some systems associated with the regulation of blood pressure. In the cardiovascular system, oxidative stress, induced by some factors such as angiotensin II (Ang II), endothelin-1, and urotensin I, promotes vascular contraction, endothelial dysfunction, and vascular remodeling, leading to vascular damage and subsequent hypertension via different mechanisms, including altering NO bioavailability or signaling, increasing intracellular calcium concentration (5, 119). In the kidney, oxidative stress leads to abnormal renal physiological functions associated with increased blood pressure via some mechanisms, including increasing renal vasoconstriction, causing dysfunction of glomerular cells and proteinuria, disrupting water and sodium homeostasis (6, 56). In the central nervous system (CNS), oxidative stress causes hypertension via some neural mechanisms, including activating the sympathetic nervous system, underpinning the processing of the cardiovascular reflexes in the medulla oblongata (65, 80). In addition, oxidative stress also induces hypertension via other mechanisms such as increasing inflammation and activating the renin/angiotensin/aldosterone system (6, 59).

The kidney is essential for the long-term regulation of arterial blood pressure. NADPH oxidase is the predominant source of ROS outside the mictochondria in the kidney (127). It is an enzyme complex comprising five subunits: membrane component protein cytochrome b588 (22^phox and gp91^phox); cytosolic components p40^phox, p47^phox, and p67^phox; and a low-molecular-weight G protein, Rac1 or Rac2. The NADPH oxidase family has seven members, Nox1, Nox2 (aka gp91^phox), Nox3, Nox4, Nox5, Duox1, and Duox. The NADPH oxidase subunits are expressed in almost all of the nephron segments, including the macula densa, mesangial cells, podocytes, and renal arterioles, including endothelial cells (92). Antioxidant enzymes, for example, SOD, the major scavenging system for removing ROS, are also well expressed in the kidney where they restrict the levels of ROS and limit hypertensive tissue injury (120, 164). Several studies have shown that excess renal production of ROS from the imbalance of pro-oxidant and antioxidant systems can initiate the development of hypertension. Renal ROS are increased in various hypertensive animal models (71, 92c, 99). Studies using several methods, including germ line gene deletion, specific RNA silencing, and pharmacological inhibitors, support the role of increased renal ROS production in the development of hypertension. For example, extracellular SOD (eSOD) gene knockout mice have high blood pressure, as well as increased renal cortical p22^phox expression, and NADPH oxidase activity (164). Specific small interfering RNA (siRNA) silencing of p22^phox in the kidney cortex is associated with reduced systemic oxidative stress and attenuation in Ang II-induced high blood pressure (103). Treatment with the NADPH oxidase inhibitor apocynin or the SOD mimetic tempol decreases systolic blood pressure, accompanied with the inhibition of renal NADPH oxidase activity in SHRs (27). Increased renal superoxide anion production is associated with the inactivation of NO, which increases afferent arteriolar tone and sodium reabsorption, and impairs tubuloglomerular feedback response, all of which are involved in the regulation of blood pressure (166). However, it should be noted that the overall effect of ROS on overall renal sodium transport is still difficult to assess because ROS can decrease or increase sodium transport, which may due to the nephron-specific effects of ROS on tubular Na⁺ transport and fluid reabsorption (111, 132, 139).

Intrarenal Dopaminergic System

Dopamine, an endogenous catecholamine, is well known as a neurotransmitter in the CNS (102). There is increasing evidence that dopamine is also an important regulator of renal function and ultimately blood pressure (14, 55). Dopamine is synthesized not only in noradrenergic and dopaminergic nerves, but also in non-neural tissues such as the kidney (141, 194, 195). Both the renal synthesis and metabolism of dopamine differ from that in neurons. Renal proximal tubules (RPTs) synthesize dopamine from the circulating or filtered dopamine precursor l-dihydroxy-phenylalanine (l-DOPA), which is then converted to dopamine by aromatic amino acid decarboxylase (AADC) (75, 141, 194, 195). Unlike neurons, RPT cells do not express dopamine β-hydroxylase, and therefore, synthesized dopamine is not metabolized to norepinephrine and epinephrine (194). Moreover, dopamine produced in the RPT is not stored, but transported across the basolateral and apical membranes and into the peritubular space and tubular lumen, respectively, where it acts on its receptors locally and in more distal nephron segments.

Dopamine, via its receptors, plays an important role in the regulation of sodium balance and blood pressure. It should be noted that the affinity of dopamine to its receptors is in the nanomolar range; higher concentrations occupy other G protein-coupled receptors (GPCRs; e.g., α-adrenergic, β-adrenergic, and serotonin receptors). Although normal circulating concentrations of dopamine (in the picomolar range) are not sufficiently high to activate dopamine receptors, high nanomolar concentrations can be attained in dopamine-producing tissues (e.g., the RPT and jejunum) (63, 194). There is evidence that intrarenal dopamine synthesis/release is modulated by alterations in dietary salt intake and intracellular sodium. Conversely, intrarenal dopamine production regulates sodium excretion. Under conditions of moderate sodium excess, locally generated dopamine acts on renal tubular and jejunal cells to decrease sodium transport. When sodium intake is enhanced, renal dopamine, acting in an autocrine/paracrine manner, is responsible for over 50% of the increase in sodium excretion (63, 187a, 194, 195).

Dysfunction of Renal Dopamine Receptors and Hypertension

The dopaminergic system exerts an autocrine/paracrine regulatory role on renal sodium transport in the kidney via its five receptor subtypes. Dopamine receptors, belonging to the α-group of the rhodopsin family of seven transmembrane GPCRs, are classified into two families: D₁-like receptors (dopamine D₁ receptor [D₁R] and dopamine D₅ receptor [D₅R]) couple to stimulatory G protein Gα_S and stimulate adenylyl cyclase (AC) activity, whereas D₂-like receptors (dopamine D₂ receptor [D₂R], dopamine D₃ receptor [D₃R], and dopamine D₄ receptor [D₄R]) couple to inhibitory G protein Gα_i/Gα_o and inhibit AC activity (63, 187a, 194, 195).

The expression of the dopamine receptor subtypes in specific nephron segments is species-dependent. For example, in humans, D₁R is expressed in all segments of the proximal tubule, thick ascending limb, distal convoluted tubule, and all segments of the collecting duct but not in the mesangial cell, podocyte, juxtaglomerular cell, or macula densa. By contrast, rats and mice express the D₁R in juxtaglomerular cell and macula densa; rats express D₁R in mesangial cell while mice express D₁R in the podocyte (63, 187a, 194, 195). No dopamine receptor is expressed in thin descending and thin ascending limb, regardless of species. Activation of renal dopamine receptors decreases renal electrolyte and water transport and is affected by many conditions, including oxidative stress (63, 187a, 194, 195). Importantly, disruption of any of the dopamine receptor genes in mice results in hypertension, the pathogenesis of which is specific for each receptor subtype, indicating the contribution of dopamine receptor on the regulation of blood pressure.

An increase in endogenous renal dopamine or the intrarenal infusion of specific dopamine receptor agonists increases sodium and water excretion; however, these effects are impaired in hypertensive states (167b, 187). Although all of the mechanisms causing the impairment are not known, impaired dopamine production or dopamine receptor dysfunction may be involved (28, 33, 48, 58, 123, 134, 136). First, an abnormal renal synthesis of dopamine may be involved in the development of hypertension (28, 33, 123, 134). Black salt-sensitive hypertensive subjects have decreased renal dopamine production (134). Caucasians with mild, untreated essential hypertension have decreased renal dopamine production, as related to the amount of sodium ingested (33). However, there are reports of increased urinary dopamine excretion in young hypertensive patients and patients with borderline hypertension, due to the elevated conversion of DOPA to dopamine in the kidney (28, 123). The reasons leading to the different results in various reports are not known. These studies indicate that age, race, and duration of hypertension affect renal dopamine production. It should be noted that the exact causal association between the hypertensive patients and abnormal dopamine metabolism is still unclear because all the above reports only observed the simple phenomenon that hypertensive patients have abnormal dopamine metabolism. Renal dopamine production is also species-dependent. For example, daily dopamine excretions are similar among Dahl salt-sensitive rats, Dahl salt-resistant rats, and Sprague-Dawley (SD) rats, but salt loading increases urinary dopamine to a greater extent in Dahl salt-sensitive and salt-resistant rats than SD rats (58). The decreased urinary dopamine in SD rats is related to a decreased conversion of DOPA to dopamine; however, urinary dopamine is similar in Dahl salt-sensitive and salt-resistant rats (58). Urinary dopamine excretion is also not different between Wistar Kyoto (WKY) rats and SHR in the basal state, and is greater in SHR than WKY rats during high-salt intake (136). Nevertheless, the natriuretic effect of D₁-like receptor agonist and the antinatriuretic effect of D₁-like receptor antagonist are impaired in the SHR (48, 68). Therefore, the renal dopamine dysfunction in hypertension starts at the receptor level. Oxidative stress may participate in the impairment of dopamine receptors (154, 175, 177). This review describes the association between dopamine receptors and oxidative stress in hypertension.

Renal D₁ receptor and hypertension

Renal-selective activation of D₁-like receptors increases sodium excretion (52). Renal-selective silencing of D₁R reduced urinary sodium excretion and urine flow in SD rats on low- or high-sodium diet, but blood pressure was not affected (162). However, the renal-selective impairment of D₁R localization in the plasma membrane with β-methyl cyclodextrin impaired the ability of the renal interstitial infusion of D₁-like receptor agonist, fenoldopam, and increased blood pressure in SD rats fed a high-sodium diet (52). Moreover, the germ line deletion of the Drd1 gene causes hypertension (4). The natriuretic effect of D₁R may be due to its ability to inhibit directly the sodium hydrogen exchanger type 3 (NHE3), sodium phosphate cotransporter type 2 (NaPi2), chloride bicarbonate (Cl⁻/HCO₃⁻) exchanger, and the epithelial sodium channel (ENaC) at the luminal membrane, as well as Na⁺-K⁺-ATPase (NKA) and the sodium bicarbonate cotransporter at the basolateral membrane (15, 23, 187a, 195). The D₁R can also indirectly exert its natriuretic effect by interacting with other hormonal factors or receptors, including Ang II, angiotensin 1–7, gastrin, Ang II types 1 and 2 receptor, and cholecystokinin B receptor (CCKBR), and other dopamine receptor subtypes (14, 30, 44, 53, 106, 110).

The D₁R function, as with other receptors, is affected by its expression in the cell membranes and compartments. For example, the D₁R is mainly located at the microvillous brush border and apical membranes in WKY rats, whereas the D₁R is mainly in the cytosol with minimal expression at the brush border and apical membrane in the SHRs. Indeed, more D₁Rs are located in endocytic vesicles in the SHR than WKY rat (183). The internalization of D₁R in renal tubules of SHR and Dahl salt-sensitive rats, may explain, in part, the diminished ability of D₁-like receptors to inhibit renal sodium transporter/channel activity in hypertension (77, 107). Impaired D₁R-mediated inhibition on sodium transports is also observed in humans with essential hypertension (108). In addition to the altered renal cellular localization of D₁R in hypertension, its expression is also decreased in obese Zucker rats, which are hypertensive (156).

Renal D₅ receptor and hypertension

The D₅R, by itself, or together with D₁R, also mediates the diuretic and natriuretic effects of D₁-like receptor; D₁R and D₅R interact in the inhibition of NHE3 and NKA activity, the D₁R primarily by cAMP, whereas the D₁R/D₅R heteromer modulates the D₁R effect through a phospholipase C pathway in RPT cells (53, 187a). The importance of the renal D₅R in the regulation of blood pressure has been proven in Drd5 knockout mice and cross-transplantation studies (13, 87). Disruption of the Drd5 gene in mice results in hypertension that is aggravated by a high-salt diet (13, 87). Blood pressure in Drd5^−/− mice transplanted with kidneys from wild-type (Drd5^+/+) mice is lower than in Drd5^−/− mice transplanted with Drd5^−/− kidneys, while blood pressure is higher in Drd5^+/+ mice transplanted with Drd5^−/− kidneys than Drd5^+/+ mice transplanted with Drd5^+/+ kidneys (13). The natriuresis in Drd5^−/− mice is decreased, which is due to the upregulation of renal sodium transporters (158). These may be partly related to the D₅R regulation of the AC isoform in lipid rafts and nonlipid rafts (186). The D₅R also interacts with Gα₁₂ and Gα₁₃, which is related to the inhibition of NHE3 activity (199). Similar to the D₁R, the natriuretic effect of D₅R may also be caused via interactions with other factors or receptors, including D₁R, angiotensin II type 1 receptor (AT₁R), and renalase (53, 54, 153).

Basal D₅R expression is decreased in RPT cells and renal brush border membranes of SHR (193). Renal D₅R expression is also decreased in obese Zucker rats (156). Moreover, the dysregulation of D₅R internalization and trafficking is involved in the higher blood pressure and blunted natriuretic response to agonist in salt-loaded mice (151). In the hypertensive state, the interactions between D₅R and other factors or receptors are disturbed, for example, activation of D₅R increased renalase protein expression and function in WKY rat RPT cells, but decreased them in SHR cells (69, 153). Such aberrant D₅R mediated-regulation may be involved in the pathogenesis of hypertension.

Renal D₂ receptor and hypertension

Activation of renal D₂R has many effects, including the inhibition of noradrenaline release from the kidney, protection against ischemia/reperfusion injury to RPTs, and maintenance of normal blood pressure (105, 117, 122). The D₂R modulates renal dopamine synthesis and protects the kidney by limiting the inflammatory reaction (84, 109). Disruption of the Drd2 gene increases blood pressure in mice that is associated with salt sensitivity, depending on the genetic background (91, 145). It should be noted that D₂R can stimulate or inhibit NKA (25, 167). The stimulatory effect of D₂-like receptors on NKA activity changes to inhibition in the presence of D₁-like receptors (79). Indeed, the natriuretic effect of dopamine depends on the activation of both D₁-like and D₂-like dopamine receptors (43, 76).

There is no difference in the renal D₂R expression and RPT cellular distribution between WKY rats and SHR. However, the D₂R is also expressed in the glomeruli in WKY rats but not SHR (131). In obese Zucker rats, renal D₂R protein expression is decreased, which induces salt sensitivity and elevates blood pressure, and can be normalized by candesartan, an AT₁R blocker (156).

Renal D₃ receptor and hypertension

Disruption of the Drd3 gene decreases urine flow rate and sodium excretion, and causes hypertension (12). However, a disruption of the Drd3 gene does not increase blood pressure regardless of salt intake in another strain of mice (135). This differential effect on blood pressure occurs despite the decreased ability of these two strains of Drd3^−/− mice to increase sodium excretion after an acute or a chronic sodium chloride load. Whether the difference in blood pressure phenotype is related to the difference in the genetic background remains to be determined. Dahl salt-resistant rats on a high-sodium diet and chronically treated with a highly selective D₃R antagonist BSF-135170 have increased blood pressure (95). Stimulation of the D₃R induces natriuresis and diuresis in Wistar rats (172). This may be partly due to the D₃R-mediated inhibition of renal NHE and NKA activity (7, 113, 114, 172). D₃R activation also modulates renal hemodynamics and tubular function, and increases amino acid-induced glomerular hyperfiltration (93, 94). In addition, stimulation of D₃R attenuates renal ischemia/reperfusion injury via increased linkage with Gα₁₂ (161).

D₃R expression is decreased in the renal cortex and RPT cell of the SHR, relative to the WKY rat (131, 188). This is reflected in in vivo studies, in which selective renal D₃R activation produces diuresis and natriuresis in the WKY rats, but not in SHRs (82, 187).

Renal D₄ receptor and hypertension

The D₄R inhibits vasopressin-dependent transepithelial sodium transport in the rat cortical collecting duct. This is secondary to the ability of D₄R to inhibit vasopressin-mediated stimulation of cAMP production (89, 138). This is exerted mainly at the basolateral membrane despite a greater expression of D₄R at the luminal membrane (124). The D₄R agonist PD168077 inhibits NKA activity in RPT cells from WKY rats but not from SHRs (140). D₄R agonist stimulation with PD168077 also decreases the stimulatory effect of insulin on NKA activity in RPT cells from WKY rats but not SHRs (198). However, renal cortical and medullary NKA activities are not altered in Drd4 knockout mice (24). The renal cortical expression of the D₄R is increased in SHRs, relative to WKY rats, although renal medullary D₄R expression is not different between these two rat strains (131). We also found that the D₄R decreases AT₁R expression in RPT cells from WKY rats but not SHRs (29). Disruption of the Drd4 gene in mice produces hypertension, which is associated with increased renal AT₁R expression (24). Thus, the D₄R, as with the other dopamine receptor subtypes, participates in the regulation of blood pressure, due, in part, to negative interaction with the AT₁R.

Disruption of the Drd4 gene does not affect body weight in mice (24). However, renal D₄R is decreased in obese Zucker rats (156). The D₄R decreases insulin receptor expression in RPT cells from WKY rats but not SHRs (198). Furthermore, the inhibition of the D₄R on AT₁R and insulin receptor is also aberrant in SHRs, which might be also involved in the pathogenesis of essential hypertension (29, 198).

Dysregulation of Dopamine Receptors in Oxidative Stress in Hypertension

D₁-like receptors

As aforementioned, the D₁-like receptor family includes at least two subtypes: D₁R and D₅R, also called D₁A receptor and D₁B receptor in rodents, which stimulate AC activity (63, 187a, 194, 195). D₁-like, not D₂-like, receptors are the major determinants of dopaminergic-mediated regulation of salt and water reabsorption induced by dopamine. Indeed, during conditions of moderate sodium balance, more than 50% of renal sodium excretion is regulated by D₁-like receptors. It should be noted that there are no agonists that can distinguish D₁R from D₅R. LE-PM436 is a selective D₅R antagonist but there is no specific D₁R agonist (53). Therefore, in our present review, the term “D₁-like receptor” is used when the study did not determine if the activation of D₁-like receptors is specifically attributable to the D₁R or D₅R.

Renal D₁R and oxidative stress

Most of studies reported that stimulation with D₁-like receptor in low concentrations reduces oxidative stress in RPT cells, not only by decreasing the generation of ROS but also by increasing their degradation (62, 88, 185). The antioxidant effect of D₁-like receptor is mainly exerted by inhibiting the pro-oxidant enzyme, NADPH oxidase, and stimulating some antioxidant enzymes such as paraoxonase 2 (PON2). Fenoldopam, a D₁-like receptor agonist, disperses Nox subunits within lipid rafts and nonlipid rafts, and decreases the expression of p22^phox and Rac1 in lipid rafts and Nox2 and Nox4 in nonlipid rafts. These actions reduce NADPH oxidase activity in human RPT cells (62). The basal NADPH oxidase activity in RPT cells is much higher in hypertensive than normotensive humans and rats but is inhibited by fenoldopam to a greater extent in the former than in the latter (88, 185). In HEK-293 cells heterologously expressing human D₁R (HEK-hD₁R), the D₁R-mediated inhibition of NADPH oxidase activity is via a protein kinase A (PKA) and protein kinase C (PKC) cross talk (184). This cross talk is due, in part, to the regulation of AC isoforms in lipid and nonlipid rafts by D₁R. D₁R, but not D₅R, increases AC activity in lipid rafts but decreases it in nonlipid rafts. Fenoldopam increases AC 5/6 isoform expression in lipid rafts but has little effect in nonlipid rafts (186).

D₁R also decreases ROS production by increasing the expression/activity of antioxidant enzymes. Short- but not long-term stimulation with fenoldopam increases the expression of PON2 in both lipid rafts and nonlipid rafts and its physical interaction with D₁R in HEK-hD₁R (173). The ability of D₁-like receptors to stimulate antioxidant enzymes, such as GSH-Px, SOD-1, and glutamylcysteine transferase, involves Nfr-2, a transcription factor, involved in antioxidant homeostasis (20). Figure 2 illustrates the effects of renal D₁R on the negative regulation of oxidative stress.

FIG. 2.

Schematic summary of the regulation of renal D₁R on oxidative stress. The broken lines indicate inhibitory effects, whereas the solid lines indicate stimulatory effects. AC, adenylyl cyclase; Co-IP, coimmunoprecipitation; D₁R, dopamine D₁ receptor; PKA, protein kinase A; PKC, protein kinase C; PON2, paraoxonase 2.

It should be noted that, although renal D₁R inhibits oxidative stress, the function of D₁R is, in turn, directly or indirectly impaired by oxidative stress. Treatment with H₂O₂ stimulates PKC leading to the activation of G protein-coupled receptor kinase 2 (GRK2), which causes serine phopshorylation of D₁R and receptor G-protein uncoupling in RPT cells, resulting in impairment in D₁-like receptor function (9). During oxidative stress, D₁R function is defective, and there is mild hypertension. However, in the presence of oxidative stress, high salt intake causes marked elevation in blood pressure in SD rats, which results from a defective renal D₁R function leading to the failure of dopamine to inhibit NKA and promote sodium excretion, indicating that oxidative stress is involved in renal D₁R dysfunction and salt-sensitive hypertension (18). Other conditions such as inflammation and hyperglycemia can also impair the function of D₁R via decreasing antioxidant enzyme and increasing oxidative stress (10, 100). However, treatment with the antioxidant tempol decreases renal D₁R hyperphosphorylation and restores D₁R-G-protein coupling and function by reducing oxidative stress in hyperglycemic rats and obese Zucker rats (10, 22). Another antioxidant sulforaphane mitigates oxidative stress, preserves D₁R function, and preventes hypertension via activation of the NRF2-phase II antioxidant enzyme pathway (20). Exercise also increases antioxidant defense and D₁R function via increasing the nuclear levels of NRF2 and decreasing oxidative stress in old rats (11).

Other mechanisms are involved in the regulation of oxidative stress on D₁R. For example, oxidative stress causes nuclear translocation of nuclear factor-κB (NF-κB) in the RPT, which contributes to defective D₁R-G-protein coupling and function via mechanisms involving PKC, membranous translocation of GRK2, and subsequent D₁R phosphorylation (16, 19). Oxidative stress also directly suppresses the transcription of D₁R gene and subsequent D₁R signaling via activation of transcription factors AP1 and SP3 (21). Both of these effects can be reversed by treatment with tempol (16, 19, 21).

Renal D₅R and oxidative stress

Compared with other dopamine receptors, the D₅R has generated significant interest for its specific characteristics. The D₅R has a higher affinity for dopamine than the D₁R and exhibits constitutive activity (139a). The D₅R also has a trafficking profile distinct from that of other dopamine receptors, which is related to the third intracellular loop of the D₅R that is necessary for PKC-mediated D₅R endocytosis. The endocytosis of D₅R, in the basal state, is dependent on clathrin and dynamin, while β-arrestin 2 is needed in the case of agonist-induced endocytosis (115, 143).

Stimulation of the D₅R results in an antioxidant response that is mediated by the inhibition of NADPH oxidase activity in the short term, and inhibition of NADPH oxidase expression, in the long term. D₅R-deficient mice are hypertensive, salt-sensitive, and in a state of systemic oxidative stress, which is related to increased renal NADPH oxidase expression and activity (177). Cross-transplantation studies showed that transplantation of a wild-type D₅R kidney into a nephrectomized Drd5^−/− mouse decreases renal Nox-2 expression. Conversely, transplantation of a Drd5^−/− kidney into a nephrectomized D₅R wild-type mouse increases renal Nox-2 expression (13). The increased blood pressure in D₅R-deficient mice may be mediated by increased ROS production because apocynin normalizes increased blood pressure. Thus, the ability of D₅R stimulation to decrease ROS production may explain, in part, the antihypertensive effect of D₅R activation (177).

The inhibition of D₅R on the NADPH oxidase activity may be partly due to D₅R-mediated inhibition of phospholipase D (PLD). PLD is an ubiquitous enzyme stimulated by many cell surface receptors that hydrolyze phospholipids to form phosphatidic acid and the free polar head group of the phospholipid substrate. PLD is involved in the pathophysiology of hypertension by increasing the formation of ROS (147, 176). Total renal PLD activity is increased in Drd5^−/− mice. PLD2, not PLD1, expression is greater in Drd5^−/− mice. Moreover, activation of the D₅R by fenoldopam in HEK-293 cells heterologously expressing human D₅R (HEK-hD₅R) cells decreases PLD2 expression and activity, indicating that the antioxidative effects of D₅R are, in part, caused by the inhibition of PLD2 (176). Dysregulation of D₅R trafficking is also associated with its impaired antioxidant function. Our studies demonstrated that the silencing of sorting nexin 1 (SNX1), involved in trafficking of internalized GPCRs, expression in RPT cells results in the failure of D₅R to internalize and bind GTP, blunting of the agonist-induced increase in cAMP production and decrease in sodium transport (151). In vivo studies showed that depletion of renal SNX1 in C57BL/6J or BALB/cJ mice increases blood pressure. The natriuretic effect of the D₁-like receptor agonist, fenoldopam, is also blunted in acutely saline-loaded BALB/cJ mice (151). Basal ROS, NOX activity, and blood pressure are higher in SNX1-deficient mice, which are normalized by apocynin, a drug that prevents NOX assembly (168).

Some protective factors against oxidative stress, such as PON2 and heme oxygenase-1 (HO-1), are also altered in the Drd5^−/− mice. Short-term stimulation with fenoldopam increases PON2 protein in nonlipid rafts, but not lipid rafts, in HEK-hD₅R; long-term fenoldopam stimulation increases PON2 protein in HEK-hD₅R, not in HEK-hD₁R cells. Moreover, renal PON2 protein is decreased in Drd5^−/− mice. In human RPT cells, silencing the Drd5 decreases PON2 expression and increases ROS production. These indicate that D₅R inhibits ROS production by regulating PON2 distribution in membrane microdomains and its renal expression (173). We also found that the activity of HO, an antioxidant enzyme, and the renal HO-1 protein expression are decreased in Drd5^−/− mice, which can be restored by the administration of hemin, an HO-1 inducer (92b). Furthermore, cellular NADPH oxidase activity is decreased by 35% in HEK-hD₅R that is abrogated with silencing of the heme oxygenase 1 (HMOX1) gene. HMOX1 siRNA also impairs the ability of fenoldopam to decrease NADPH oxidase activity in HEK-hD₅R cells (92b). These indicate that the high blood pressure and increased ROS production in Drd5^−/− mice may be caused by decreased HO-1 expression and activity.

D₅R single-nucleotide polymorphisms (SNPs) are reported to be involved in oxidative stress in hypertension. Humans carry SNPs in the DRD5 gene, some of which confer diminished D₅R function and abnormal coupling with AC (67). The human D₅R173F>L (hD₅R^173F>L) mutation markedly impairs stimulation of cAMP production (92a, 154). On normal salt diet, the blood pressure, renal NADPH oxidase activity, and AT₁R expression are higher in hD₅R^173F>L than hD₅R^WT transgenic mice. However, after 2 weeks on high-salt diet, the blood pressure and renal NADPH oxidase activity, but not AT₁R expression, are increased in hD₅R^173F>L but not in hD₅R^WT transgenic mice. These can be reversed by the AT₁R antagonist candesartan, indicating that the ability of the hD₅R to negatively regulate the renal NADPH oxidase activity and AT₁R function may have important implications in the pathogenesis of salt-sensitive blood pressure (92a). Our other studies also demonstrated the role of thioredoxin 1 (Trx1), an antioxidant enzyme, in the impaired sodium excretion of hD₅R^173F>L transgenic mice (154). hD₅R^173F>L transgenic mice have higher blood pressure, lower basal urine flow and sodium excretion, and impaired agonist (fenoldopam)-mediated natriuresis and diuresis, which are due, in part, to decreased renal Trx1 expression and increased ROS production. Hyperphosphorylation of hD₅R^173F>L, with its dissociation from Gα_s and Gα_q, is the key factor in the impaired renal D₅R function and increased blood pressure of hD₅R^173F>L mice (154). Figure 3 illustrates the effects of renal D₅R on the regulation of oxidative stress.

FIG. 3.

Schematic view of effects of renal D₅R on the regulation of oxidative stress. The broken lines indicate inhibitory effects, whereas the solid lines indicate stimulatory effects. D₅R, dopamine D₅ receptor; HO-1, heme oxygenase-1; PLD2, phospholipase D2; Trx1, thioredoxin 1.

D₂-like receptors

Renal D₂R and oxidative stress

Decreasing oxidative stress is one of the mechanisms by which D₂R regulates blood pressure. Disruption of Drd2, the D₂R gene, increases ROS production and causes ROS-dependent hypertension (8). Drd2^−/− mice have increased urinary excretion of 8-isoprostane, a product of lipid peroxidation, NADPH activity, and expression of Nox1, Nox2, and Nox4, but decreased expression of the antioxidant enzyme heme oxygenase-2 (HO-2) in the kidney (8, 36). Apocynin, an inhibitor of NADPH oxidase, normalizes the blood pressure in Drd2^−/− mice (8). Thus, the D₂R negatively regulates ROS production by inhibiting pro-oxidant but enabling antioxidant systems (8, 36). The D₂R in the adrenal gland is also implicated in aldosterone regulation because urinary aldosterone is increased in Drd2^−/− mice. However, spirolactone, a competitive antagonist of the mineralocorticoid receptor, only normalized the renal expression of Nox1 and Nox4, but had no effects on the excretion of 8-isoprostane and the expression of Nox2 or HO-2, indicating that the increased ROS production in Drd2^−/− mice is only partially mediated by impaired aldosterone regulation (8). Similar to the results in Drd2 knockout mice, silencing the D₂R in mouse RPT cells increases ROS production (36). The results from the germ line deletion of Drd2 or silencing of Drd2 in mouse RPT cells are supported by studies stimulating the D₂R. Dopamine and D₂R agonists have free radical scavenging and antioxidant activities. Stimulation of D₂R in mouse or human RPT cells decreases hyperoxidized peroxiredoxins and reduces ROS production, accompanied by a decrease in Nox-4 expression and NADPH oxidase activity (36, 175).

The antioxidant effect of D₂R involves its interaction with other proteins, including PON2, DJ-1 (also known as Park 7), and sestrin2 (36, 174, 175). PON2, which belongs to the paraoxonase gene family, is expressed in the kidney, acting to protect against cellular oxidative stress. Renal PON2 is involved in the inhibition of renal NADPH oxidase activity and ROS production and contributes to the maintenance of normal blood pressure. Our studies found that D₂R colocalizes with PON2 in the brush border of mouse RPTs; both D₂R and PON2 are found in nonlipid rafts and lipid rafts and physically interacts with each other (175). Renal PON2 protein is decreased in Drd2^−/− mice. Treatment of human RPT cells with the D₂R agonist quinpirole decreases ROS production that is related to D₂R agonist stimulation of PON2 (175). These indicate that PON2 is positively regulated by D₂R and may, in part, mediate the inhibitory effect of renal D₂R on NADPH oxidase activity and ROS production.

The D₂R also interacts with another antioxidant protein DJ-1. DJ-1 is a multifunctional oxidative stress response protein that defends cells against ROS and mitochondrial damage (36, 40). D₂R and DJ-1 are expressed in the mouse kidney and colocalize and coimmunoprecipitate in mouse RPT cells. Mice with germ line deletion of Drd2 or with renal-selective Drd2 silencing are hypertensive and have increased ROS production and renal cortical expression of NOX4, but decreased expression of DJ-1. Conversely, treatment of mouse RPT cells with a D₂R agonist increases DJ-1 expression and decreases NOX4 expression and NADPH oxidase activity. Renal-selective silencing of DJ-1 in mice increases blood pressure, renal Nox4 expression, and NADPH oxidase activity (36). Silencing DJ-1 or Drd2 in mouse RPT cells and kidneys also decreases Nrf2 expression and activity and increases ROS production (35). These studies suggest that the inhibitory effect of D₂R on renal ROS production is at least, in part, mediated by the positive regulation of DJ-1.

Sestrin2, another antioxidant protein, is involved in the effect of renal D₂R on blood pressure. We found that the antioxidant action of D₂R is mediated, in part, by a positive regulation of sestrin2 expression. Drd2 germ line deletion in mice or Drd2 silencing in human RPT cells decreases, whereas D₂R stimulation increases sestrin2 expression (174). Sestrin2 decreases ROS production and prevents cellular damage from oxidative stress by catalyzing the reduction of hyperoxidized peroxiredoxins. D₂R-induced sestrin2 activation is dependent on PON2, and depletion of PON2 increases peroxiredoxin hyperoxidation and sestrin2 degradation (174). In vivo renal selective silencing of sestrin2 in mice increases renal oxidative stress and blood pressure (174). These results suggest that the D₂R, via PON2/DJ-1/sestrin2 pathways, keeps a normal renal redox balance, which contributes to the maintenance of normal blood pressure.

D₂R-mediated inhibition on oxidative stress may also be associated with decreasing inflammation. Oxidative stress is positively linked to inflammation and vice versa. D₂R protects the kidney by limiting the inflammatory reaction; impaired D₂R function results in renal inflammation and damage (196). The decrease or lack of D₂R function enhances Akt phosphorylation by decreasing the expression of the protein phosphatase 2A (PP2A) regulatory subunit PPP2R2C. This activates NF-κB, a proinflammatory transcription factor that can be repressed or activated by oxidative stress and vice versa (167a, 197). However, renal-selective D₂R rescue by the retrograde renal infusion of adeno-associated virus (AAV) vector carrying D₂R reduces the expression of proinflammatory factors and kidney injury, preserves renal function, and normalizes the blood pressure (81). Moreover, RPT cells of subjects bearing D₂R SNPs (rs6276, rs6277, and rs1800497), via the miR-217/Wnt5a/Ror2 pathway, decrease D₂R expression and express a proinflammatory phenotype with increased TNF-α, cytokines, and chemokines and a profibrotic phenotype with increased transforming growth factor-beta 1 (TGF-β₁) (61, 74), which are associated with increased ROS production. However, the mechanisms involved in the renal D₂R-mediated inhibition of inflammation and oxidative stress need to be determined. Figure 4 illustrates the effects of renal D₂R on the regulation of oxidative stress.

FIG. 4.

Schematic presentation of the role of renal D₂R on the regulation of oxidative stress. The broken lines indicate inhibitory effects, whereas the solid lines indicate stimulatory effects. ALDO, aldosterone; D₂R, dopamine D₂ receptor; HO-2, heme oxygenase-2; NF-κB, nuclear factor-κB; TGF-β₁, transforming growth factor-beta 1.

Renal D₃R/D₄R and oxidative stress

Everett and Senogles reported that the D₃R, via a complex with RhoA, stimulates PLD activity in human HEK293 cells overexpressing the rat D₃R (45). However, in rat nerve cells (oligodendrocytes), the D₃R has antioxidant effects. D₃R activation protects oligodendrocyte injury caused by glutamate oxidative stress (121); pramipexole, a D₂R/D₃R agonist but with a much higher affinity to the D₃R (pK_i = 8.4–8.7) than D₂R (pK_i = 5.1–7.4), protects against free radical-mediated lipid peroxidation in dopaminergic neurons (200). The hypertension in Drd3^−/− mice is mild and not associated with increased ROS production, but may be due to the compensatory mechanism that occurs with deletion of the Drd3 gene. For example, disruption of the Drd3 gene in mice leads to the increased expression of D₅R (155), which has antioxidant activity (13, 177). In addition, activation of D₃R increases D₁R, D₅R, and endothelin B receptor expression and their mediated inhibitory effect on NKA activity in RPT cells; all of these receptors have antioxidant activity (69, 182, 192). It is not known if endothelin B receptor expression is increased in the kidney of Drd3^−/− mice, but D₁R expression is not. Whether or not D₃R has a direct antioxidant activity remains to be determined.

Similar to the renal D₃R, whether the D₄R has an antioxidative effect in the kidney or not is still unknown. However, D₄R exerts an antioxidant effect in the nervous system (70, 130). We can speculate that renal D₄R may have an indirect antioxidative activity in the kidney. The D₄R can negatively regulate the renal and vascular expressions of AT₁R and insulin receptor (29, 198), which are pro-oxidants (83, 180). Activation of AT₁R increases D₄R expression in rat RPT cells and renal AT₁R expression is increased in Drd4^−/− mice (24, 140). As with the D₃R, a direct antioxidant effect of D₄R remains to be determined.

Aberrant Regulation of Renal Dopamine Receptors on Oxidative Stress in Hypertensive Status

Oxidative stress, whether systemic or renal, is present in animal models of hypertension and human essential hypertension (3, 6). The inhibitory effect of dopamine receptors on oxidative stress is impaired in hypertensive animal models. The phosphorylation of renal dopamine receptors is increased and persistent in various hypertensive animal models. This may be the main reason for the impaired renal dopamine receptor-mediated inhibition on oxidative stress in hypertensive status (2, 32, 92c, 146, 157, 181). For example, the long-term exposure of pregnant SD rats to fine particulate matter induces hypertension accompanied by persistent phosphorylation of the renal D₁R. The D₁R antioxidant effect is decreased, resulting in an increased ROS production (92c). In the hypertensive offspring of these rats, the inhibition of ROS production by tempol, an oxygen free radical scavenger, reduces blood pressure and increases sodium excretion, accompanied by a normalization of D₁R function that is related to phosphorylation (181). Indeed, the hyperphosphorylation and dysfunction of renal D₁R and increased ROS, found in hypertensive offspring, induced by adverse maternal exposures, such as maternal diabetes mellitus, fine particulate matter, and lipopolysaccharide, are normalized by tempol (96, 157, 181).

In obese Zucker rats, which manifest with moderate degree of hypertension, both hyperglycemia and hyperinsulinism, via the upregulation of mitogen-activated protein kinase (MAPK), reduce renal D₁R affinity and G protein coupling, leading to increased oxidative stress, which can be ameliorated by antioxidants (17). In the other hypertensive animal models such as old age-associated hypertension or hyperinsulinemia-induced hypertension, increased oxidative stress has also been shown to contribute to the hyperphosphorylation and defective function of renal D₁R, which can be restored by antioxidants (2, 32, 146). The age-associated hypertension may be due to the different mechanisms that affect the renal D₁R response to high-salt diet. In FBN rats, high-salt diet increases renal D₁R expression in adult rats, but decreases D₁R expression and increases renal gp91^phox and NHE3 expression in aged rats (116).

Abnormal intracellular localization of the dopamine receptor may be also involved in the impaired renal dopamine receptor-mediated inhibition of oxidative stress in hypertensive states. For example, D₁R expression is decreased at the RPT cell surface membrane, accompanied by increased expression in early or sorting endosome, in SHRs, relative to WKY rats. These findings indicate that in the SHR, the majority of D₁Rs in RPTs are internalized and fail to be recycled to the cell surface membrane, resulting in defective D₁R function (183). Additional factors, including GPCRs, may be also involved in the impaired renal dopamine receptor-mediated inhibition of oxidative stress in hypertension. For example, stimulation of CCKBR increases the cell membrane expression of the D₁R in WKY rat RPT cells, but not in SHRs (30). Short-term Ang II treatment decreases cell surface membrane D₁R protein in WKY rats but not in SHR RPT cells. Moreover, D₁R and AT₁R colocalization and coimmunoprecipitation are greater in WKY rats than in SHRs (191). The dopamine receptor exerts its physiological functions, including antioxidant effects, via its proper internalization and reinsertion to the plasma membrane after activation with its agonist. Thus, the abnormal intracellular localization of dopamine receptor causes inactivation, impaired reactivation, and function, and results in increased ROS production in hypertension.

Mechanisms involved in the abnormal regulation of renal dopamine receptors on oxidative stress

There are several mechanisms involved in the abnormal regulation of renal dopamine receptors, resulting in oxidative stress. G protein-coupled receptor kinase 4 (GRK4), a subtype in a family of seven serine/threonine protein kinases characterized by their ability to specifically recognize and phosphorylate agonist-activated GPCRs, plays a crucial role in the regulation of renal D₁R phosphorylation and cellular trafficking (169, 170). Increased GRK4 activity due to increased expression or GRK4 gene variants impairs renal D₁R function in hypertension. Basal renal GRK4 and serine-phosphorylated D₁R levels are higher in SHRs than WKY rats. The renal-selective silencing of GRK4 decreases the quantity of serine-phosphorylated D₁R and increases urine flow and sodium excretion to a greater extent in SHRs than WKY rats (125).

The impaired renal dopamine receptor-mediated inhibition on oxidative stress in hypertension may be due to the phosphorylation of D₁R because the D₁R expression is not changed, but its phosphorylation is increased in hypertension, which is associated with the increased activity of GRK2 and GRK4 (47, 78, 157). Studies of others and ours have also shown that the increased expression and activity of GRK4 enhance D₁R phosphorylation, decrease D₁R function, leading to the increased ROS production. These effects can be reversed by antioxidants in some hypertensive animal models, including age-related hypertension, and hypertensive offspring induced by adverse maternal exposures such as fine particulate matter and lipopolysaccharide (2, 32, 92c, 157, 181).

GRK4 gene variants, including GRK4 486V, 142V, and 65L, play important roles in the pathogenesis of hypertension and may provide novel therapeutic antihypertensive strategies (149, 170, 190). Decreasing renal GRK4 expression attenuates the increase in blood pressure with age and decreases the protein excretion in SHRs (125). Indeed, expressing human (h) GRK4γ142A>V but not the human GRK4γ (hGRK4γ) gene in transgenic mice produces hypertension and impairs the diuretic and natriuretic effects of D₁-like agonist stimulation (47). Our meta-analysis revealed a significant association of GRK4 486V with hypertension, with an odds ratio of 1.5 (95% confidence interval: 1.2–1.9) (190). Depending on the genetic background of the mouse, hGRK4γ wild type prevents salt-sensitive hypertension, whereas hGRK4γ 486V converts a salt-resistant phenotype to a salt-sensitive phenotype (160). The elevated blood pressure of hGRK4γ142A>V transgenic mice is not associated with increased ROS production (159). By contrast, salt-sensitive hGRK4γ486A>V transgenic mice have increased renal oxidative stress (41). The hypertension of salt-sensitive hGRK4γ486V transgenic mice can be partially prevented by chronic tempol treatment (41). Figure 5 illustrates the regulation of D₁R by GRK4 in the kidney. In addition, we have also reported that GRK4 regulates the phosphorylation and function of the D₃R in human proximal tubule cells (152). However, the role of this regulation in the modulation of renal oxidative stress needs further study. Moreover, the role of GRK4 on the other renal dopamine receptors still remains unclear.

FIG. 5.

Regulation of D₁R by GRK4 in the kidney. The increased expression and activity of GRK4 and its GRK4 gene variants (R65L, A142V, and A486V) cause D₁R phosphorylation and D₁R/G protein uncoupling, followed with inactivity of D₁R and increased the activity of NADPH oxidases, leading to the increased ROS production and blood pressure. The broken lines indicate inhibitory effects, whereas the solid lines indicate stimulatory effects. D₁R, dopamine receptor type 1; GRK4, G protein-coupled receptor kinase 4.

GRK2, another subtype of G protein-coupled receptor kinase, also phosphorylates D₁R and uncouples D₁R from signaling proteins. Increased expression of GRK2 also impairs D₁R function that can result in increased oxidative stress and hypertension, including age-associated hypertension, hyperinsulinemia-mediated hypertension, and hypertensive offspring induced by adverse maternal exposures such as lipopolysaccharide and maternal diabetes mellitus (15, 16, 46, 96, 157). As with GRK4-related hypertension, GRK2-related hypertension can be normalized by the antioxidants (46).

The abnormal cellular trafficking of dopamine receptors may be another important mechanism involved in the abnormal regulation of renal dopamine receptors that can result in oxidative stress and hypertension. The SNX family consists of a diverse group of cytoplasmic and membrane-associated proteins orchestrating the process of cargo sorting, through the endosomal network (171). We reported that sorting nexin 5 (SNX5), a component of the mammalian retromer, is critical to D₁R internalization and trafficking to the plasma membrane (150). However, renal SNX5 expression is decreased in SHRs and RPT cells from hypertensive humans, which causes aberrant D₁R trafficking and impaired D₁R function, including D₁R-mediated antioxidative effects and sodium excretion (86, 150). Depletion of renal SNX5 expression in SHRs causes a further increase in systolic blood pressure and a decrease in sodium excretion (150). Moreover, renal-selective silencing of Snx5 in C57Bl/6J mice increases blood insulin and glucose levels, decreases urinary insulin excretion, and causes insulin resistance, which are associated with impaired D₁R function (86, 150). Insulin can increase the production of ROS.

Another SNX subtype, SNX1, the first mammalian SNX, is essential for the trafficking and function of D₅R. Our studies showed that renal-selective silencing of SNX1 in C57Bl/6 and BALB/c mice impairs the agonist-activated D₅R internalization, prevents GTP binding, blunts cAMP response, impairs natriuresis, and increases blood pressure (151). Moreover, germ line deletion of the Snx1 gene in mice increases blood pressure and enhances ROS production that may be related to an increase in the expression of NADPH oxidase components in the kidney. The oxidative stress associated with germ line deletion of the Snx1 gene in mice is due to the impaired D₅R antioxidative function and normalized by the antioxidant apocynin (168). These studies indicate that abnormal regulation of the trafficking of dopamine receptors caused by decreased abundance of SNXs leads to impaired receptor function, increased oxidative stress, and hypertension. The targeting SNXs may represent a novel mechanism for the treatment of essential hypertension.

Conclusions

In summary, increasing evidence shows that renal dopamine receptors exert their antioxidative activity in the kidney and play an important role in the regulation of sodium excretion and blood pressure (Table 1). The aberration of such regulation may be involved in the pathogenesis of hypertension. Increased understanding of the role of reciprocal regulations between renal dopamine receptors and oxidative stress in the regulation of renal function and blood pressure may give us a novel insight into the pathogenesis of hypertension.

Table 1.

Summary of Renal Dopamine Receptors, Oxidative Stress, and Hypertension

Receptor subtype	Effects of renal dopamine receptors on the physiological status	Functional deficits in hypertension	Manifestation in gene knockout/knockdown mice or gene mutant	References
D1R	Inhibits renal sodium transport; induces natriuresis and diuresis; disperses NOX subunits, decreases NADPH oxidase expression and activity; interacts with antioxidant enzymes such as PON2.	Decreased D1R-mediated natriuresis and diuresis in SHR and Dahl salt-sensitive rats; impaired D1R-mediated inhibition of sodium transport in hypertensive subjects; reduced D1R expression in obese Zucker rats; increased renal NADPH oxidase expression and activity in hypertensive rats and subjects.	Increased blood pressure after selective inhibition of the renal dopamine subtype D1R with AS-ODN in conscious SD rats; reduced urinary sodium excretion and urine output and elevated blood pressure in Drd1−/− mice.	(4, 15, 23, 63, 77, 88, 107, 108, 133, 156, 162, 173, 184)
D5R	Inhibits renal sodium transport; induces natriuresis and diuresis; inhibits NADPH oxidase expression and activity; increases PON2 expression.	Decreased D5R expression in RPT cells and renal brush border membranes of SHR; impaired D5R internalization and trafficking in salt-induced hypertension.	Elevated blood pressure, and increased expression of renal sodium transporters in Drd5−/− mice; enhanced systemic oxidative stress in Drd5−/− mice; increased renal PLD activity, NADPH oxidase expression and activity, decreased renal expression of PON2 and HO-1 protein, and impaired HO activity in Drd5−/− mice; increased blood pressure and renal NADPH oxidase activity, and decreased urine sodium excretion in hD5R173F>L transgenic mice.	(13, 73, 87, 92b, 151, 154, 158, 173, 176, 177, 193)
D2R	Inhibits noradrenaline release from the kidney; modulates renal local dopamine synthesis; limits renal inflammation; inhibits renal NKA activity in the presence of D1-like receptors; decreases hyperoxidized peroxiredoxins and reduces ROS production, accompanied with decreased NOX4 expression and NADPH oxidase activity; interacts with antioxidant enzymes, including PON2, DJ-1, and sestrin2.	Decreased D2R expression in obese Zucker rats.	Hypertension associated with salt sensitivity, and sodium retention in Drd2−/− mice; increased renal ROS production, NADPH oxidase expression and activity, and renal inflammation in Drd2−/− mice; renal inflammation and fibrosis induced by D2R SNPs (rs6276, rs6277, and rs1800497).	(8, 36, 61, 74, 79, 81, 84, 109, 122, 145, 156, 174, 175, 196)
D3R	Induces natriuresis and diuresis; inhibits renal NHE and NKA activity; modulates renal hemodynamics and tubular function, and increases amino acid-induced glomerular hyperfiltration.	Decreased renal D3R expression in the cortex and RPT cells of SHR; impaired D3R-imediated diuresis and natriuresis in SHR; impaired inhibition of sodium transport in SHRs and in hypertensive Dahl salt-sensitive rats.	Hypertension and decreased urine flow rate and sodium excretion in Drd3−/− mice.	(7, 12, 93–95, 113, 114, 131, 155, 172, 187, 188)
D4R	Inhibits vasopressin-dependent transepithelial Na+ transport and osmotic water permeability; inhibits NKA activity.	Impaired D4R-imediated NKA inhibiton in RPT cells of SHR; decreased D4R expression in obese Zucker rats.	Hypertension associated with increased renal AT1R expression in Drd4−/− mice.	(24, 138, 140, 156)

AS-ODN, antisense oligodeoxynucleotide; AT₁R, angiotensin II type 1 receptor; D₁R, dopamine D₁ receptor; D₂R, dopamine D₂ receptor; D₃R, dopamine D₃ receptor; D₄R, dopamine D₄ receptor; D₅R, dopamine D₅ receptor; HO-1, heme oxygenase-1; NADPH, nicotinamide-adenine dinucleotide phosphate; NHE, sodium hydrogen exchanger; NKA, Na⁺-K⁺-ATPase; PLD, phospholipase D; PON2, paraoxonase 2; ROS, reactive oxygen species; RPT, renal proximal tubule; SD, Sprague-Dawley; SHR, spontaneously hypertensive rat; SNP, single-nucleotide polymorphism.

However, there are still some questions that need to be studied in the future, for example, the exact causal relationship between increased oxidative stress and the impaired renal dopamine receptors is still unclear; the direct antioxidant activity of renal D₃R and D₄R still needs to be proved; more details of mechanisms involved in the abnormal regulation of renal dopamine receptors on oxidative stress are needed to be determined. Dopamine receptor-mediated inhibition of oxidative stress may be found to play a potential role in the antihypertensive treatment, which may represent a valuable pharmaceutical target for potential therapeutic approaches in the treatment of hypertension.

Abbreviations Used

AC

adenylyl cyclase

Ang II

angiotensin II

AT₁R

angiotensin II type 1 receptor

CCKBR

cholecystokinin B receptor

CNS

central nervous system

D₁R

dopamine D₁ receptor

D₂R

dopamine D₂ receptor

D₃R

dopamine D₃ receptor

D₄R

dopamine D₄ receptor

D₅R

dopamine D₅ receptor

GPCRs

G protein-coupled receptors

GR

glutathione reductase

GRK2

G protein-coupled receptor kinase 2

GRK4

G protein-coupled receptor kinase 4

GSH-Px

glutathione peroxidase

GST

glutathione transferase

H₂O₂

hydrogen peroxide

HEK-hD₁R

HEK-293 cells heterologously expressing human D₁R

HEK-hD₅R

HEK-293 cells heterologously expressing human D₅R

hGRK4γ

human GRK4γ

HMOX1

heme oxygenase gene

HO-1

heme oxygenase-1

HO-2

heme oxygenase-2

l-DOPA

l-dihydroxy-phenylalanine

NADPH

nicotinamide-adenine dinucleotide phosphate

NF-κB

nuclear factor-κB

NHE3

sodium hydrogen exchanger type 3

NKA

Na⁺-K⁺-ATPase

NO

nitric oxide

PKC

protein kinase C

PLD

phospholipase D

PON2

paraoxonase 2

ROS

reactive oxygen species

RPTs

renal proximal tubules

SD

Sprague-Dawley

SHR

spontaneously hypertensive rat

siRNA

small interfering RNA

SNP

single-nucleotide polymorphism

SNX1

sorting nexin 1

SNX5

sorting nexin 5

SOD

superoxide dismutase

Trx1

thioredoxin 1

WKY

Wistar Kyoto

Funding Information

These studies were supported, in part, by grants from the National Key R&D Program of China (2018YFC1312700), the National Natural Science Foundation of China (31730043, 81770425), Program of Innovative Research Team by National Natural Science Foundation (81721001), Key Program of The Third Affiliated Hospital of Chongqing Medical University (KY19024), the National Institutes of Health (R01-DK039308, P01-HL074940), and the Clinical Medical Research Talent Training Program of The Third Military Medical University (2018XLC10I2).