Interaction between Gα12 and Gα13 protein subunits and dopamine receptors in renal proximal tubules

The cells of the renal proximal tubules (RPTs) have high aromatic L-amino-acid decarboxylase activity. Filtered or circulating L-3,4-dihydroxyphenylalanine can be converted to dopamine after uptake into this extraneuronal compartment, without being subsequently converted to norepinephrine.¹ Peripheral dopamine has been characterized as an important modulator of both renal sodium excretion and blood pressure by acting directly on renal epithelial ion transport and by modulating the secretion/release of other hormonal/humoral molecules. These hormonal and humoral molecules include aldosterone, catecholamines, renin and vasopressin, each of which contribute to the regulation of sodium homeostasis and blood pressure. In addition, other hormones may interact with dopamine produced in RPTs to increase (for example, atrial natriuretic peptide) or decrease (for example, angiotensin) its inhibitory effect on tubular sodium reabsorption. The actions of endogenous renal dopamine on water and electrolyte transport are modest under euvolemic conditions, but become magnified during moderate sodium excess. Thus, following a moderate acute or chronic sodium load, up to 50% of sodium excretion is mediated by dopamine produced by the RPTs.²

The natriuretic effect of peripheral dopamine is exerted by two major receptor classes, D₁-like and D₂-like receptors, which belong to the rhodopsin-like family of membrane receptors called G-protein-coupled receptors (GPCRs). GPCRs have specific resultant actions due to their heterotrimeric G-protein subunits composed of α, β and γ.^{2, 3} D₁-like receptors include D₁ and D₅ subtypes, which stimulate adenylyl cyclase. D₁ couples to Gαs and Gαq, whereas D₅ couples to Gαs, Gα12 and Gα13.^{4, 5, 6} The linkage of G-protein subunits to the specific D₁-like receptor is tissue specific. D₂-like receptors include D₂, D₃ and D₄ subtypes, which are coupled to Gαi. Gαi subunits inhibit adenylyl cyclase and calcium channel activities.^{2, 7} In RPTs, D₁, D₃, D₄ and D₅ receptors are expressed. Although the quantitative contribution of a particular dopamine receptor subtype to renal sodium transport in RPTs has not been studied previously, most in vivo studies suggested that dopamine-induced natriuresis is mediated principally by D₁-like receptor subtypes. In fact, a number of studies evidenced that the activation of D₁-like receptors in RPTs decreases sodium reabsorption by the inhibition of both the Na⁺/HCO₃⁻ co-transporter and Na⁺-K⁺-ATPase activities in the basolateral membranes. Moreover, D₁-like receptors in RPTs demonstrate inhibition of the Na⁺-H⁺ antiporter (NHE3), the Na⁺-Pi co-transporter and the Cl⁻/HCO₃⁻ antiporter in the apical membranes. Evidence has been provided suggesting that the dopamine-induced natriuretic response resulting from activation of tubular D₁-like receptors is diminished in both spontaneously hypertensive rats and in humans with essential hypertension. This compromised natriuretic response in hypertension was described to result from alterations occurring at the receptor level as well as at the cellular signaling pathway level, which ultimately decreases tubular sodium reabsorption.⁸

The effects of the D₂-like receptors, independent of the D₁-like receptors, on sodium excretion were not consistent, ranging from natriuresis to antinatriuresis and no effect. This lack of consistency was attributed to the use of drugs with poor D₂-like receptor subtype selectivity. However, the current perception is that D₂-like receptors may function synergistically with D₁-like receptors in RPTs, where they may potentiate the inhibitory effects of D₁-like receptors on NHE3, the Na⁺-Pi co-transporter and Na⁺-K⁺-ATPase activities.⁹ In the rat kidney, the major D₂-like receptor in RPTs is the D₃ receptor. The mechanisms underlying the interaction between D₃ and D₁ receptors were recently investigated using immortalized RPT cells.¹⁰ In these studies, the D₃ receptor agonist PD128907 increased the immunoreactive expression of the D₁ receptors in a concentration-dependent and time-dependent manner. These data suggest synergism between D₃ and D₁ receptors capable of acutely increasing sodium excretion. In addition, co-immunoprecipitation of the D₃ and D₁ receptors in RPT cells was observed.¹⁰ Together, these results indicate that the natriuretic effects of D₃ receptor activation in RPT cells could be due, at least in part, to D₃ receptor-mediated increases in D₁ receptor expression, specifically, total and cell surface membrane expression. Additionally, these results indicate direct D₃ and D₁ receptor interaction. The interaction between D₃ and D₁ receptors was impaired in RPT cells from spontaneously hypertensive rats, which provided evidence favoring their combined contribution to compromised sodium excretion and increased blood pressure in this rat hypertension model.¹⁰

In this issue of Hypertension Research, Zhang et al.¹¹ report on their findings that D₃ receptors in RPTs could bind to the fourth family member of the G-protein subunit (Gα12 and Gα13) when activated by the D₃ receptor-selective agonist PD128907. This binding was accompanied by the co-localization and co-immunoprecipitation of the D₃ receptor and Gα12 and Gα13 in renal brush border membranes and RPT cells. The compound PD128907 inhibited the Na⁺-K⁺-ATPase in RPTs in a concentration-dependent manner. Gα12 and Gα13 are known to stimulate sodium reabsorption in RPTs by increasing pump and transporter activity (more specifically, Na⁺-K⁺-ATPase and NHE3). Therefore, these results indicate that the association of the D₃ receptors with Gα12 and Gα13 may be one of the mechanisms underlying the natriuretic effect induced by stimulation of the D₃ receptors in RPT cells.

As previously mentioned, Gα12 and Gα13 were already linked to the D₅ receptor, but not to the D₁ receptors.⁶ A linkage to the D₅ receptors was found in RPTs in native kidneys, in immortalized RPT cells and in HEK293 cells heterologously expressing the D₅ receptor. Laser confocal microscopy revealed the co-localization of the D₅ receptor with Gα12 and Gα13 at the brush border membranes and subjacent areas. In these elegant experiments, the authors also provided evidence that the D₁-like agonist fenoldopam increased the interaction between the D₅ receptor with both Gα12 and Gα13 in brush border membranes.⁶ These results, when viewed together with the findings reported by Zhang et al.¹¹ in this issue, indicate that Gα12 and Gα13 may represent a common intracellular pathway of D₁-like (D₅) and D₂-like (D₃) receptors in RPT cells (see Figure 1). Because D₁-like and D₂-like receptors may function synergistically in RPT cells, it would be interesting to examine the influence of D₁ stimulation on the co-localization of D₃ receptors with Gα12 and Gα13. In addition, as the interaction between D₃ and D₁ receptors is impaired in RPT cells from spontaneously hypertensive rats, it would be interesting to examine the linkage between D₃ receptors and Gα12 and Gα13 in RPT cells in this rat hypertension model.

Figure 1

The interaction of Gα12 and Gα13 with dopamine receptors in the renal proximal tubules (RPTs). While experiencing conditions of moderately increased NaCl intake, the renal D₁, D₃ and D₅ receptors are stimulated by dopamine in the kidney (1). D₃ receptor stimulation increases the immunoreactive expression of D₁ receptors (2). Stimulation of the D₃ receptors is accompanied by increased co-immunoprecipitation and internalization of Gα12 and Gα13 with the D₃ receptors (3). Stimulation of the D₅ receptors is accompanied by co-localization and immunoprecipitation of Gα12 and Gα13 with the D₅ receptors (4). D₁, D₃ and D₅ receptor activation decreases sodium reabsorption and contributes to blood pressure control (5). It is suggested that both the D₁-like (D₅) and D₂-like (D₃) receptors may participate in the regulation of sodium transport by hampering Gα12 and Gα13 actions because Gα12 and Gα13 stimulate sodium transport by modulating the activities of the Na⁺-K⁺-ATPase and NHE3 in RPTs.

Gα12 and Gα13 are expressed in other locations besides brush border membranes and subjacent areas to RPT cells. Gα12 is expressed in the ascending limb of the loop of Henle and cortical collecting ducts. Gα13 is expressed in the distal tubules, the medullary collecting duct and the juxtaglomerular apparatus.⁶ Therefore, the results by Zhang et al.¹¹ reinforce the view that each of the dopamine receptor subtypes—alone or by interacting with the other dopamine receptor subtypes, other GPCRs and G-protein subunits—regulate tubular sodium transport uniquely. Accordingly, the ultimate natriuretic effect of dopamine will be the sum of the interactions among the D₁-like and D₂-like dopamine receptors, other GPCRs, such as endothelin and angiotensin receptors, and G-protein subunits. Knowledge of the regulatory pathways involving differential G-protein subunit linkages on different dopamine receptors may provide new approaches to the pharmacological regulation of sodium excretion and blood pressure control.