Ligand-induced modification of a surface cAMP receptor of Dictyostelium discoideum does not require its occupancy

In Dictyostelium discoideum amoebae, cAMP-induced phosphorylation of the surface cAMP receptor is associated with a discrete transition in its electrophoretic mobility. The native and modified forms of the receptor are designated R and D (Mr = 40,000 and 43,000). The relationship of the number of receptors which are modified as a function of the receptors which bind cAMP was investigated. Modification was assessed by determining the amounts of R and D forms in Western blots which detect all receptors whether or not they are exposed on the surface. Cyclic AMP or the analog, adenosine 3',5'-monophosphorothioate ((Rp)-cAMPS), induced a loss of cAMP-binding activity (down-regulation), which was not accompanied by a loss of the receptor protein. About 60% of the receptors do not bind cAMP in the absence of Ca2+ and are unmasked by 10 mM Ca2+. However, the fraction of receptors which are modified in response to cAMP is equal in the absence or presence of Ca2+. (Rp)-cAMPs induces down-regulation (50%) but not modification. Addition of cAMP, following down-regulation by (Rp)-cAMPS, causes all receptors to be modified. cAMP induces both down-regulation (80%) and modification. Modification is more readily reversed than down-regulation: 30 min after removal of cAMP, receptors remain down-regulated (57%) but are found in the R form. All receptors shift to the D form when cAMP is readded to the cells. These results indicate that exposed, as well as cryptic and down-regulated receptors, are modified in response to the cAMP stimulus.     

Berberine Reduces Fibronectin Expression by Suppressing the S1P-S1P2 Receptor Pathway in Experimental Diabetic Nephropathy Models

The accumulation of glomerular extracellular matrix (ECM) is one of the critical pathological characteristics of diabetic renal fibrosis. Fibronectin (FN) is an important constituent of ECM. Our previous studies indicate that the activation of the sphingosine kinase 1 (SphK1)-sphingosine 1- phosphate (S1P) signaling pathway plays a key regulatory role in FN production in glomerular mesangial cells (GMCs) under diabetic condition. Among the five S1P receptors, the activation of S1P2 receptor is the most abundant. Berberine (BBR) treatment also effectively inhibits SphK1 activity and S1P production in the kidneys of diabetic models, thus improving renal injury. Based on these data, we further explored whether BBR could prevent FN production in GMCs under diabetic condition via the S1P2 receptor. Here, we showed that BBR significantly down-regulated the expression of S1P2 receptor in diabetic rat kidneys and GMCs exposed to high glucose (HG) and simultaneously inhibited S1P2 receptor-mediated FN overproduction. Further, BBR also obviously suppressed the activation of NF-κB induced by HG, which was accompanied by reduced S1P2 receptor and FN expression. Taken together, our findings suggest that BBR reduces FN expression by acting on the S1P2 receptor in the mesangium under diabetic condition. The role of BBR in S1P2 receptor expression regulation could closely associate with its inhibitory effect on NF-κB activation.     

Alterations in atrial natriuretic peptide and its receptors in streptozotocin-induced diabetic rat kidneys

In this study the effect of diabetes mellitus on atrial natriuretic peptide (ANP) receptors in streptozotocin- (STZ-) induced diabetic rat kidneys was studied. Moreover, plasma ANP concentration was evaluated in diabetic and control rats by using radioimmunoassay. In addition, the expression of ANP in the kidneys of control and diabetic rats was evaluated by immunohistochemistry. Body-weight loss and increased glucose levels were used as indices of diabetes mellitus in the STZ-induced rats. There was a significant loss in the body weight of the diabetic rats compared to controls. The efficacy of STZ administration was confirmed by rising blood glucose levels, which were significantly higher in diabetic rats compared to controls. Plasma ANP concentration was significantly greater in the diabetic rats in comparison with controls. Moreover, our immunohistochemical results show that the expression of ANP in diabetic rats was higher than that in age-matched controls. ANP was observed in the cells lining the proximal convoluted tubules in the cortex. The distribution and levels of ANP receptors in the kidneys of diabetic rats and age-matched controls were investigated using quantitative receptor autoradiography. Our results demonstrate significant decrease in ANP receptors in the kidneys of the diabetic rats compared to controls. The significant decrease was found in the juxtaglomerular medulla, inner medulla, and the papillae. The decrease in ANP receptors observed in the diabetic kidneys could have pathological consequences resulting in renal resistance to ANP in diabetes.     

Involvement of AT(1) angiotensin receptors in the vasomodulatory effect of des-aspartate-angiotensin I in the rat renal vasculature

Angiotensin II is known to act primarily on the angiotensin AT(1) receptors to mediate its physiological and pathological actions. Des-aspartate-angiotensin I (DAA-I) is a bioactive angiotensin peptide and have been shown to have contrasting vascular actions to angiotensin II. Previous work in this laboratory has demonstrated an overwhelming vasodepressor modulation on angiotensin II-induced vasoconstriction by DAA-I. The present study investigated the involvement of the AT(1) receptor in the actions of DAA-I on angiotensin II-induced vascular actions in the renal vasculature of normotensive Wistar-Kyoto rats (WKY), spontaneously hypertensive rats (SHR) and streptozotocin (STZ)-induced diabetic rats. The findings revealed that the angiotensin receptor in rat kidney homogenate was mainly of the AT(1) subtype. The AT(1) receptor density was significantly higher in the kidney of the SHR. The increase in AT(1) receptor density was also confirmed by RT-PCR and Western blot analysis. In contrast, AT(1) receptor density was significantly reduced in the kidney of the streptozotocin-induced diabetic rat. Perfusion with 10(-9)M DAA-I reduced the AT(1) receptor density in the kidneys of WKY and SHR rats suggesting that the previously observed vasodepressor modulation of the nonapeptide could be due to down-regulation or internalization of AT(1) receptors. RT-PCR and Western blot analysis showed no significant changes in the content of AT(1) receptor mRNA and protein. This supports the suggestion that DAA-I causes internalization of AT(1) receptors. In the streptozotocin-induced diabetic rat, no significant changes in renal AT(1) receptor density and expression were seen when its kidneys were similarly perfused with DAA-I.     

Alterations in atrial natriuretic peptide and its receptors in streptozotocin-induced diabetic rat kidneys

In this study the effect of diabetes mellitus on atrial natriuretic peptide (ANP) receptors in streptozotocin- (STZ-) induced diabetic rat kidneys was studied. Moreover, plasma ANP concentration was evaluated in diabetic and control rats by using radioimmunoassay. In addition, the expression of ANP in the kidneys of control and diabetic rats was evaluated by immunohistochemistry. Body-weight loss and increased glucose levels were used as indices of diabetes mellitus in the STZ-induced rats. There was a significant loss in the body weight of the diabetic rats compared to controls. The efficacy of STZ administration was confirmed by rising blood glucose levels, which were significantly higher in diabetic rats compared to controls. Plasma ANP concentration was significantly greater in the diabetic rats in comparison with controls. Moreover, our immunohistochemical results show that the expression of ANP in diabetic rats was higher than that in age-matched controls. ANP was observed in the cells lining the proximal convoluted tubules in the cortex. The distribution and levels of ANP receptors in the kidneys of diabetic rats and age-matched controls were investigated using quantitative receptor autoradiography. Our results demonstrate significant decrease in ANP receptors in the kidneys of the diabetic rats compared to controls. The significant decrease was found in the juxtaglomerular medulla, inner medulla, and the papillae. The decrease in ANP receptors observed in the diabetic kidneys could have pathological consequences resulting in renal resistance to ANP in diabetes. (Mol Cell Biochem 261: 3–8, 2004)     

Molecular signaling mechanisms of renal gluconeogenesis in nondiabetic and diabetic conditions

The kidneys are as involved as the liver in gluconeogenesis which can significantly contribute to hyperglycemia in the diabetic condition. Substantial evidence has demonstrated the overexpression of rate-limiting gluconeogenic enzymes, especially phosphoenolpyruvate carboxykinase and glucose 6 phosphatase, and the accelerated glucose release both in the isolated proximal tubular cells and in the kidneys of diabetic animal models and diabetic patients. The aim of this review is to provide an insight into the mechanisms that accelerate renal gluconeogenesis in the diabetic conditions and the therapeutic approaches that could affect this process in the kidney. Increase in gluconeogenic substrates, reduced insulin concentration or insulin resistance, downregulation of insulin receptors and insulin signaling, oxidative stress, and inappropriate activation of the renin–angiotensin system are likely to participate in enhancing renal gluconeogenesis in the diabetic milieu. Several studies have suggested that controlling glucose metabolism at the renal level favors effective overall glycemic control in both type 1 and type 2 diabetes. Therefore, renal gluconeogenesis may be a promising target for effective glycemic control as a therapeutic strategy in diabetes.     

The Altered Renal and Hepatic Expression of Solute Carrier Transporters (SLCs) in Type 1 Diabetic Mice

Diabetes mellitus is a chronic metabolic disorder that significantly affects human health and well-being. The Solute carrier transporters (SLCs), particularly the Organic anion/cation transporters (Oats/Octs/Octns), Organic anion transporting polypeptides (Oatps) and Oligopeptide transporters (Pepts) are essential membrane proteins responsible for cellular uptake of many endogenous and exogenous substances such as clinically important drugs. They are widely expressed in mammalian key organs especially the kidney and liver, in which they facilitate the influx of various drug molecules, thereby determining their distribution and elimination in body. The altered expression of SLCs in diabetes mellitus could have a profound and clinically significant influence on drug therapies. In this study, we extensively investigated the renal and hepatic expression of twenty essential SLCs in the type 1 diabetic Ins2^Akita murine model that develops both hyperglycemia and diabetes-related complications using real-time PCR and immunoblotting analysis. We found that the renal expression of mOatp1a1, mOatp1a6, mOat1, mOat3, mOat5, mOct2 and mPept2 was decreased; while that of mPept1 was increased at the mRNA level in the diabetic mice compared with non-diabetic controls. We found up-regulated mRNA expression of mOatp1a4, mOatp1c1, mOctn2, mOct3 and mPept1 as well as down-regulation of mOatp1a1 in the livers of diabetic mice. We confirmed the altered protein expression of several SLCs in diabetic mice, especially the decreased renal and hepatic expression of mOatp1a1. We also found down-regulated protein expression of mOat3 and mOctn1 in the kidneys as well as increased protein expression of mOatp1a4 and mOct3 in the livers of diabetic mice. Our findings contribute to better understanding the modulation of SLC transporters in type 1 diabetes mellitus, which is likely to affect the pharmacokinetic performance of drugs that are transported by these transporters and therefore, forms the basis of future therapeutic optimization of regimens in patients with type 1 diabetes mellitus.     

Tissue-specific expression of PPAR mRNAs in diabetic rats and divergent effects of cilostazol

Cilostazol and ligands of peroxisome proliferator-activated receptors (PPARs) have been effectively used to alleviate diabetic complications, but the common and tissue-specific expression patterns of PPARs in different tissues in diabetic patients and those treated with cilostazol have not been reported. Here, we aimed to assess the effects of diabetes and cilostazol on mRNA expression of PPARalpha and PPARgamma in the aorta, renal cortex, and retina of diabetic rats treated with cilostazol for 8 weeks. PPARalpha mRNA expression showed uniform downregulation in all these tissues in diabetic rats, and this effect was reversed by cilostazol treatment. Surprisingly, PPARgamma mRNA expression was reduced in the renal cortex and retina, yet increased in the aorta of diabetic rats, although cilostazol still reversed these changes. Interestingly, cilostazol, a well-known phosphodiesterase 3 inhibitor and cAMP elevator, augmented cAMP content only in the aorta, but showed no significant effects in the renal cortex of diabetic rats. In conclusion, mRNA expression of PPARs is tissue-specific in diabetes and may be differently affected by cilostazol, possibly because of its tissue-specific effects on cAMP content.     

Extracellular cAMP inhibits D1 dopamine receptor expression in CAD catecholaminergic cells via A2a adenosine receptors

The expression of D1 dopamine (DA) receptor gene is regulated during development, aging, and pathophysiology. The extracellular factors and signaling mechanisms that modulate the expression of D1 DA receptor have not been well characterized. Here, we present novel evidence that endogenous D1 DA receptor expression is inhibited by extracellular cAMP in the Cath.A Derived (CAD) catecholaminergic neuronal cell line. CAD cells express the multi-drug resistance protein 5 transporters and secrete cAMP. Addition of exogenous cAMP decreases D1 receptor mRNA and protein greater than fourfold in 24 h. The cAMP-induced decrease of D1 receptor mRNA levels is blocked by cGMP and by 1,3-dipropyl-8-(p-sulfo-phenyl)xanthine, an inhibitor of ecto-phosphodiestrase. Extracellular AMP, a metabolite of cAMP, also independently decreased D1 receptor mRNA levels. Inhibitors of ecto-nucleotidases, alpha,beta-methyleneadenosine 5'-di-phosphate and GMP, completely blocked the decrease of D1 receptor mRNA by extracellular cAMP, but only partially blocked the decrease induced by extracellular AMP. Levamisole, an inhibitor of tissue non-specific alkaline phosphatase, completely blocked the AMP-induced decrease of D1 receptor mRNA. The extracellular cAMP, AMP, and adenosine (ADO)-induced decrease in D1 receptor mRNA expression are mediated by A2a ADO receptor subtype. The results suggest a novel molecular mechanism linking activation of A2a ADO receptors with inhibition of D1 DA receptor expression.     

Reduced Adenosine Uptake and Its Contribution to Signaling that Mediates Profibrotic Activation in Renal Tubular Epithelial Cells: Implication in Diabetic Nephropathy

Altered nucleoside levels may be linked to pathogenic signaling through adenosine receptors. We hypothesized that adenosine dysregulation contributes to fibrosis in diabetic kidney disease. Our findings indicate that high glucose levels and experimental diabetes decreased uptake activity through the equilibrative nucleoside transporter 1 (ENT1) in proximal tubule cells. In addition, a correlation between increased plasma content of adenosine and a marker of renal fibrosis in diabetic rats was evidenced. At the cellular level, exposure of HK2 cells to high glucose, TGF-β and the general adenosine receptor agonist NECA, induced the expression of profibrotic cell activation markers α-SMA and fibronectin. These effects can be avoided by using a selective antagonist of the adenosine A₃ receptor subtype in vitro. Furthermore, induction of fibrosis marker α-SMA was prevented by the A₃ receptor antagonist in diabetic rat kidneys. In conclusion, we evidenced the contribution of purinergic signaling to renal fibrosis in experimental diabetic nephropathy.     

Modification of adenosine A1 and A2A receptor density in the hippocampus of streptozotocin-induced diabetic rats

Adenosine A(1) and A(2A) receptors are neuromodulatory systems that can control mnemonic behavior, which is modified by diabetes. Since the density of these adenosine receptors can change upon chronic noxious brain conditions, we now tested if the density of A(1) and A(2A) receptors was modified in the hippocampus of streptozotocin-induced diabetic rats. The binding density of the selective A(1) receptor antagonist, (3)H-DPCPX, was decreased by 36% in total hippocampal membranes 7 days after induction of diabetes and this down-regulation was maintained after 30 and 90 days, which was also confirmed by Western blot analysis of A(1) receptor immunoreactivity. In contrast, the binding density of the selective A(2A) receptor antagonist, (3)H-SCH 58261, was enhanced by 83% in total hippocampal membranes 7 days after induction of diabetes and this up-regulation persisted after 30 and 90 days. These results show that the balance between inhibitory A(1) and facilitatory A(2A) adenosine receptors is modified in the hippocampus of streptozotocin-induced diabetic rats. Thus, the most abundant A(1) receptors are down-regulated and there is an up-regulation of A(2A) receptors, suggesting a gain of function of hippocampal A(2A) receptors compared to A(1) receptors in diabetes, as has been observed in other chronic noxious brain conditions where A(2A) receptor blockade affords robust neuroprotection.     

Nox4 NAD(P)H oxidase mediates hypertrophy and fibronectin expression in the diabetic kidney

Renal hypertrophy and extracellular matrix accumulation are early features of diabetic nephropathy. We investigated the role of the NAD(P)H oxidase Nox4 in generation of reactive oxygen species (ROS), hypertrophy, and fibronectin expression in a rat model of type 1 diabetes induced by streptozotocin. Phosphorothioated antisense (AS) or sense oligonucleotides for Nox4 were administered for 2 weeks with an osmotic minipump 72 h after streptozotocin treatment. Nox4 protein expression was increased in diabetic kidney cortex compared with non-diabetic controls and was down-regulated in AS-treated animals. AS oligonucleotides inhibited NADPH-dependent ROS generation in renal cortical and glomerular homogenates. ROS generation by intact isolated glomeruli from diabetic animals was increased compared with glomeruli isolated from AS-treated animals. AS treatment reduced whole kidney and glomerular hypertrophy. Moreover, the increased expression of fibronectin protein was markedly reduced in renal cortex including glomeruli of AS-treated diabetic rats. Akt/protein kinase B and ERK1/2, two protein kinases critical for cell growth and hypertrophy, were activated in diabetes, and AS treatment almost abolished their activation. In cultured mesangial cells, high glucose increased NADPH oxidase activity and fibronectin expression, effects that were prevented in cells transfected with AS oligonucleotides. These data establish a role for Nox4 as the major source of ROS in the kidneys during early stages of diabetes and establish that Nox4-derived ROS mediate renal hypertrophy and increased fibronectin expression.     

Prostaglandin E2 down-regulation of cytochrome P-450 2B1 expression induced by phenobarbital is through EP2 receptor in rat hepatocytes

Cytochrome P-450 is an important bioactivation-detoxification system in vivo. Its expression is regulated by foreign chemicals and dietary factors, and lipids have been found to regulate its gene expression. We showed previously that prostaglandin E(2) (PGE(2)), a fatty acid metabolite, down-regulates cytochrome P-450 2B1 (CYP 2B1) expression induced by phenobarbital. The objective of the present study was to determine whether PGE(2) type 2 receptor (EP(2))-which is coupled to Gs-protein when bound by PGE(2), leading to cAMP production-is involved in this down-regulation. We also determined the possible roles of EP(2) downstream pathways in this down-regulation. We used a primary rat hepatocyte culture model in which EP(2) was shown to be present to study this question. The intracellular cAMP concentration in primary rat hepatocytes was significantly higher after treatment with 1microM PGE(2) than after treatment with 0, 0.01, or 0.1microM PGE(2). Butaprost, an EP(2) agonist, down-regulated CYP 2B1 expression in a dose-dependent manner. SQ22536, an adenylate cyclase inhibitor, reversed the down-regulation by PGE(2) as did H-89, a protein kinase A inhibitor. These results suggest that EP(2) and the downstream pathways of cAMP and protein kinase A are involved in the down-regulation of CYP 2B1 expression by PGE(2) in the presence of phenobarbital.     

Effect of diabetes on natriuresis in the presence of ouabain and ethacrynic acid in perfused rat kidney

1. The effect of diabetes on renal sodium retention was investigated. 2. The technique involved retrograde perfusion from the renal veins via the kidneys, and then through the renal arteries and dorsal aorta. 3. Sodium retention by diabetic rat kidney was 58% lower than that in the normal rats. 4. Ouabain (15 mM) in perfusate increased sodium retention by 30% in normal rat kidney as compared to a 54% increase in diabetic rat kidney. 5. Ethacrynic acid (1 mM) in perfusate resulted in a 42% reduction in sodium retention in the normal rat kidney as compared to a 43% decrease in the diabetic rat kidney. 6. Control of hyperglycemia in diabetic rats with insulin therapy resulted in sodium retention that is not significantly different from that of normal rats. 7. The results suggest that diabetes has no effect on the peritubular ouabain-sensitive Na--K-ATPase pump, or the luminal ethacrynic acid-sensitive Na-K counter transport pump. Furthermore, the data suggest a reversible effect of diabetes on sodium retention during insulin therapy.     

The diabetic rat kidney mediates inosituria and selective urinary partitioning of D-chiro-inositol

Diabetic nephropathy is a serious complication of diabetes mellitus with a pressing need for effective metabolic markers to detect renal impairment. Of potential significance are the inositol compounds, myo-inositol (MI), and the less abundant stereoisomer, D-chiro-inositol (DCI), which are excreted at increased levels in the urine in diabetes mellitus, a phenomenon known as inosituria. There is also a selective urinary excretion of DCI compared to MI. As the biological origins of altered inositol metabolism in diabetes mellitus are unknown, the aim of this study was to determine whether the diabetic kidney was directly responsible. Kidneys isolated from four-week streptozotocin-induced diabetic rats were characterized by a 3-fold reduction in glomerular filtration rate (GFR) compared to matched non-diabetic kidneys. When perfused with fixed quantities of MI (50 µM) and DCI (5 µM) under normoglycemic conditions (5 mM glucose), GFR-normalized urinary excretion of MI was increased by 1.7-fold in diabetic vs. non-diabetic kidneys. By comparison, GFR-normalized urinary excretion of DCI was increased by 4-fold. Perfusion conditions replicating hyperglycemia (20 mM glucose) potentiated DCI but not MI urinary excretion in both non-diabetic and diabetic kidneys. Overall, there was a 2.4-fold increase in DCI urinary excretion compared to MI in diabetic kidneys that was independent of glucose ambience. This increased urinary excretion of DCI and MI in diabetic kidneys occurred despite increased renal expression of the inositol transporters, sodium myo-inositol transporter subtype 1 and 2 (SMIT1 and SMIT2). These findings show that the diabetic kidney primarily mediates inosituria and altered urinary partitioning of MI and DCI. Urinary inositol levels might therefore serve as an indicator of impaired renal function in diabetes mellitus with wider implications for monitoring chronic kidney disease.     

Endothelin antagonism prevents early EGFR transactivation but not increased matrix metalloproteinase activity in diabetes

Although past studies have demonstrated decreased renal matrix metalloproteinase (MMP) activity in type 1 diabetes and in mesangial cells grown under high glucose conditions, renal MMP expression and activity in type 2 diabetes and the regulation of MMPs by profibrotic factors involved in diabetic renal complications such as endothelin-1 (ET-1) remained unknown. The renal expression and activity of MMPs in type 2 diabetic Goto-Kakizaki (GK) rats treated with vehicle or ET(A) receptor selective antagonist ABT-627 for 4 wk were assessed by gelatin zymography, fluorogenic gelatinase assay, and immunoblotting. In addition, expression and phosphorylation of epidermal growth factor receptor (EGFR) and connective tissue growth factor were evaluated by immunoblotting. Renal sections stained with Masson trichrome were used to investigate kidney structure. MMP-2 activity and protein levels were significantly increased in both cortical and medullary regions in the GK rats. Membrane-bound MMP (MT1-MMP), MMP-9, and fibronectin levels were also increased, and ABT-627 treatment did not have an effect on MMP activity and expression. Histological analysis of kidneys did not reveal any structural changes. Phosphorylation of EGFR was significantly increased in the diabetic animals, and ABT-627 treatment prevented this increase, suggesting ET-1-mediated transactivation of EGFR. These results suggest that there is early upregulation of renal MMPs in the absence of any kidney damage. Although the ET(A) receptor subtype is not involved in the early activation of MMPs in type 2 diabetes, ET-1 contributes to transactivation of growth-promoting and profibrotic EGFR.     

Hydrogen peroxide causes uncoupling of dopamine D1-like receptors from G proteins via a mechanism involving protein kinase C and G-protein-coupled receptor kinase 2

Dopamine, via activation of D1-like receptors, inhibits Na,K-ATPase and Na,H-exchanger in renal proximal tubules and promotes sodium excretion. This effect of dopamine is not seen in conditions associated with oxidative stress such as hypertension, diabetes, and aging due to uncoupling of D1-like receptors from G proteins. To identify the role of oxidative stress in uncoupling of the D1-like receptors, we utilized primary cultures from rat renal proximal tubules. Hydrogen peroxide (H2O2), an oxidant, treatment to the cell cultures increased the level of malondialdehyde, a marker of oxidative damage. Further, H2O2 decreased membranous D1-like receptor numbers and proteins, D1-like agonist (SKF 38393)-mediated [35S]GTPgammaS binding and SKF 38393-mediated inhibition of Na,K-ATPase. Moreover, H2O2 treatment to the cultures caused membranous translocation of G-protein-coupled receptor kinase 2 (GRK 2) and increased serine phosphorylation of D1A receptors accompanied by an increase in protein kinase C (PKC) activity. Interestingly, PKC inhibitors blocked the H2O2-mediated stimulation of GRK 2 and serine phosphorylation of D1A receptors. Further, GRK 2 antisense but not scrambled oligonucleotides attenuated the effect of H2O2 on membranous expression of GRK 2. Moreover, direct activation of PKC with phorbol ester (PMA) resulted in reduction of SKF 38393-mediated [35S]GTPgammaS binding. We conclude that H2O2 stimulates PKC leading to the activation of GRK 2, which causes serine phopshorylation of D1A receptors and receptor G-protein uncoupling in these cells, resulting in impairment in D1-like receptor function.