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R A Felder

Publications and source records attributed to R A Felder.

At least 73 records · Page 4Linked to original sources
PubMed1993

Renal dopamine-1 receptors in hypertensive inbred rat strains with and without hyperactivity.

Renal dopamine-1 (DA-1) receptors are involved in the regulation of sodium transport in several nephron segments, including the proximal convoluted tubule (PCT). DA-1 receptors in the PCT and cortical collecting duct of normotensive rats are linked to the stimulation of adenylyl cyclase (AC). We have reported a defect in the DA-1 receptor/AC coupling in the PCT of the spontaneously hypertensive rat (SHR) of the Okamoto-Aoki strain. Hyperactivity and hypertension are both expressed in the SHR. To determine if the DA-1 receptor coupling defect is associated with hyperactivity or hypertension, we studied the DA-1 receptor in the PCT of two new inbred rat strains derived from the SHR: the hyperactive WKHA and the hypertensive WKHT rat. Tail-cuff blood pressures taken at 4 weeks indicated that WKHT rats were not hypertensive (86 +/- 3 mm Hg, n = 6), whereas at 12 weeks systolic pressures in both SHR and WKHT rats exceeded 150 mm Hg. Hyperactivity, however, was noted in WKHA rats even at this early age. Basal AC activity was similar in WKHA and WKHT PCT in either age group. In the older rats, the DA-1 agonist fenoldopam (10(-7) mol/L) stimulated AC activity in WKHA (70.6 +/- 16.1 fmol per 3 mm PCT per 20 minutes, n = 3) but not in WKHT PCT (43.3 +/- 5.3 fmol per 3 mm PCT per 20 minutes, n = 4). Gpp(NH)p (10(-5) mol/L), a nonhydrolyzable GTP analogue, stimulated AC activity to a similar extent in WKHA and WKHT PCT.(ABSTRACT TRUNCATED AT 250 WORDS)

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1993

D1A dopamine receptor stimulation inhibits Na+/K(+)-ATPase activity through protein kinase A.

We stably expressed the rat D1A dopamine receptor in mouse fibroblast LTK- cells and obtained specific ligand binding and functional activity characteristic of the D1A dopamine receptor coupled to stimulation of adenylyl cyclase. In the transfected cells, the selective D1 agonist fenoldopam caused a concentration-dependent inhibition of Na+/K(+)-ATPase activity, achieving maximum inhibition of approximately 30%. The latter was abolished by the selective D1 antagonist (+)-SCH 23390 and by the specific protein kinase A inhibitor protein kinase inhibitor-(6-22) amide. In the nontransfected cells, fenoldopam did not affect Na+/K(+)-ATPase activity. 8-Chlorophenylthio-cAMP inhibited Na+/K(+)-ATPase activity in both transfected and nontransfected cells; this effect was blocked by protein kinase inhibitor-(6-22). These results indicate that the inhibition of Na+/K(+)-ATPase activity induced by agonist occupancy of D1A receptors is mediated by protein kinase A.

Adenylyl Cyclases↗
PubMed1993

Robotic chromatography: development and evaluation of automated instrumentation for assay of glycohemoglobin.

The measurement of glycohemoglobin (GHb) by boronate affinity chromatography is useful in monitoring long-term glucose control in diabetic subjects. The inherent disadvantage of this method is the hands-on time required because the hemoglobin fractions are separated on individual disposable columns. To overcome this disadvantage, we have programmed a Hamilton Microlab 2200 automated pipetting cartesian robot to complete the procedure, from the aspiration of blood from the sample-collection tube to the transfer of the separated hemoglobin fractions to a microtiter plate for absorbance measurement. This automated robotic system can analyze 96 specimens, including patients' samples and control material, in approximately 3 h. The precision (CV) of the method ranged from 1.6% to 3.5% within-run and from 2.7% to 3.5% day-to-day. The results correlated with those obtained with the Accuflex semiautomated robot, which used the identical disposable column, and those obtained with a Primus high-performance liquid chromatograph, which used a regenerated microparticle column. Automation of the GHb procedure allowed improved throughput, reduced labor cost, improved precision, and offered greater laboratory safety.

Autoanalysis↗
PubMed1992

Intrarenal DA2 dopamine receptor stimulation in the conscious dog.

DA2 dopamine receptors are present in renal blood vessels and glomeruli. Stimulation of DA1 dopamine receptors leads to renal vasodilation, diuresis, and natriuresis, but a functional role for renal DA2 receptors is largely unknown. We investigated the possible role of DA2 receptors in the control of renal function by intrarenal infusion of a highly specific DA2 agonist, LY 171555 (LY), in conscious uninephrectomized dogs (n = 5) in metabolic balance at sodium intake of 40 meq/day. The infusion of LY at 0.5 pmol.kg-1.min-1 did not change the urinary sodium excretion or renal hemodynamic function. A significant dose-dependent antidiuresis (F = 8.1, P less than 0.0001) and antinatriuresis (F = 93.3, P less than 0.0001) and a decrease in filtration fraction (F = 2.3, P less than 0.02) occurred as the LY dose was increased from 1.0 to 10.0 pmol.kg-1.min-1. There were no changes in systemic plasma renin activity, plasma aldosterone concentration, or mean arterial pressure during intrarenal LY administration. These data suggest that intrarenal DA2 receptor stimulation with LY decreases renal sodium excretion in part by hemodynamic mechanisms. Renal dopamine may act at vascular and/or glomerular DA2 receptors to modulate renal function.

Animals↗
PubMed1992

Renal dopamine receptors and pre- and post-cAMP-mediated Na+ transport defect in spontaneously hypertensive rats.

We have reported defective coupling of the renal tubular DA1 dopamine receptor to adenylyl cyclase in both the spontaneously hypertensive rat (SHR) and the Dahl salt-sensitive rat. Since Na+, 5'-guanyl imidodiphosphate [Gpp(NH)p], and N-ethylmaleimide (NEM) reduce agonist affinity for brain D1 dopamine receptors, we compared the effects of these agents on agonist affinity in proximal tubules from SHR and its normotensive control, the Wistar-Kyoto rat (WKY), to delineate further the site of the DA1-adenylyl cyclase coupling defect. In WKY, the D1/DA1 agonist, fenoldopam, competed for 125I-Sch 23982 at a high-affinity site (KiH = 1.8 +/- 0.8 x 10(-8) M) and a low-affinity site (KiL = 7.6 +/- 1.1 x 10(-5) M, n = 6). Na+ (150 mM) or Gpp(NH)p (10(-4) M) converted KiH to KiL. NEM, which alkylates sulfhydryl groups, also converted all the binding to KiL; this effect could be prevented by prior treatment with 10(-4) M fenoldopam. In contrast, in SHR, fenoldopam detected only a KiL (7.8 +/- 1.4 x 10(-5) M, n = 6). Neither Na+, Gpp(NH)p, nor NEM had any effect on KiL. To study a functional expression of these binding sites, the effect of 5 x 10(-5) M fenoldopam or 8-(chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (8-CPT-cAMP) on Na+/H+ exchange activity in proximal tubular brush-border membrane vesicles was tested. In WKY, the inhibitory effects of these agents on the exchanger increased with the age of the rat.(ABSTRACT TRUNCATED AT 250 WORDS)

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1992

Persistent defective coupling of dopamine-1 receptors to G proteins after solubilization from kidney proximal tubules of hypertensive rats.

The natriuretic effect of dopamine-1 (DA-1) agonists is reduced in spontaneously hypertensive rat (SHR), partly because of defective DA-1 receptor-adenylate cyclase (AC) coupling in renal proximal convoluted tubules. To investigate this defective coupling, DA-1 dopamine receptors from renal proximal tubules were solubilized and reconstituted into phospholipid vesicles. The binding of DA-1-selective ligand [125I]SCH 23982 was specific and saturable, with no differences in receptor density or Kd between SHR and normotensive rats (Wistar-Kyoto rats; WKY). Competition experiments of the reconstituted DA-1 dopamine receptors in WKY with a DA-1-selective agonist, SKF R-38393, revealed the presence of high- (Kh = 350 +/- 209 nM) and low-affinity (Kl = 70,500 +/- 39,500 nM) binding sites. 100 microM Gpp(NH)p abolished the agonist high-affinity sites, converting them to a low-affinity state (Ki = 33,650 +/- 10,850 nM). In SHR, one affinity site was noted (Ki = 13,800 +/- 500) and was not modulated by Gpp(NH)p (Ki = 11,505 +/- 2,295). The absence of guanine nucleotide-sensitive agonist high-affinity sites may explain the defective DA-1/AC coupling mechanism in the SHR.

Animals↗
PubMed1992

The renal dopamine receptors.

Dopamine is an endogenous catecholamine that modulates many functions including behavior, movement, nerve conduction, hormone synthesis and release, blood pressure, and ion fluxes. Dopamine receptors in the brain have been classically divided into D1 and D2 subtypes, based on pharmacological data. However, molecular biology techniques have identified many more dopamine receptor subtypes. Several of the receptors cloned from the brain correspond to the classically described D1 and D2 receptors. Several D1 receptor subtypes have been cloned (D1A, D1B, and D5) and are each coupled to the stimulation of adenylyl cyclase. The D2 receptor has two isoforms, a shorter form, composed of 415 amino acids, is termed the D2short receptor. The long form, called the D2long receptor, is composed of 444 amino acids; both are coupled to the inhibition of adenylyl cyclase. The D3 and D4 receptors are closely related to, but clearly distinct from, the D2 receptor. They have not yet been linked to adenylyl cyclase activity. Outside of the central nervous system, the peripheral dopamine receptors have been classified into the DA1 and DA2 subtypes, on the basis of synaptic localization. The pharmacological properties of DA1 receptors roughly approximate those of D1 and D5 receptors, whereas those of DA2 receptors approximate those of D2 receptors. A renal dopamine receptor with some pharmacological features of the D2 receptor but not linked to adenylyl cyclase has been described in the renal cortex and inner medulla. In the inner medulla, this D2-like receptor, termed DA2k, is linked to stimulation of prostaglandin E2 production, apparently due to stimulation of phospholipase A2. Of the cloned dopamine receptors, only the mRNA of the D3 receptor has been reported in the kidney. The DA1 receptor in the kidney is associated with renal vasodilation and an increase in electrolyte excretion. The DA1-related vasodilation and inhibition of electrolyte transport is mediated by cAMP. The role of renal DA2 receptors remains to be clarified. Although DA1 and DA2 receptors may act in concert to decrease transport in the renal proximal convoluted tubule, the overall function of DA2 receptors may be actually the opposite of those noted for DA1 receptors. Dopamine has been postulated to act as an intrarenal natriuretic hormone. Moreover, an aberrant renal dopaminergic system may play a role in the pathogenesis of some forms of hypertension. A decreased renal production of dopamine and/or a defective transduction of the dopamine signal is/are present in some animal models of experimental hypertension as well as in some forms of human essential hypertension.

Humans↗
PubMed1992

Robotic automation of cyclosporine analysis in whole blood.

Cyclosporin A (CsA) is currently the most selective immunosuppressant used clinically to prevent organ rejection after transplantation. We used a reprogrammable robot, the Benchmate (Zymark Inc., Hopkinton, MA), to automate much of the sample preparation involved in solid-phase extraction of CsA. Lysed whole-blood specimens containing previously added internal standard were placed on the Benchmate in the specimen-holding area, and a C18 Bond-Elut column was placed in the top of the sample tube. Specimen extraction from this point was handled automatically by the Benchmate, after which we manually injected the sample into the HPLC system for quantification. Analytical recovery of CsA in two concentrations of calibrator was similar for the manual and Benchmate methods. Analytical imprecision for the Benchmate was less than for manual extraction: within-run imprecision (CV) was less than 8% at either concentration. Between-run imprecision, determined by having the Benchmate extract CsA from two specimens each day for 20 days, was less than 9%. Patients' specimens were extracted manually (x) or by using the Benchmate robot (y); the results, compared by linear regression, agreed well: (y = 0.99 (SD 0.02)x + 2.6 (SD 2.8) micrograms/L (Sy[x = 7.59 micrograms/L, n = 27). We conclude that the Benchmate usefully reduces the manual steps necessary to extract specimens before HPLC analysis for CsA.

Autoanalysis↗
PubMed1991

Prospective evaluation of maternal serum human chorionic gonadotropin levels in 3428 pregnancies.

As part of a multicenter prospective study, second-trimester human chorionic gonadotropin and alpha-fetoprotein concentrations were evaluated. Data included maternal age, human chorionic gonadotropin level, alpha-fetoprotein level, weight, race, and pregnancy outcome of 3428 pregnancies at between 15 and 20 weeks' gestation. The results of the study indicate that human chorionic gonadotropin levels decrease as maternal weight increases, that weight-adjusted human chorionic gonadotropin levels for Oriental and black women are higher than for white or Hispanic women, and that twin pregnancies have higher human chorionic gonadotropin levels than singleton pregnancies. Of 255 pregnancies that did not have normal outcomes, 54 (21.2%) had human chorionic gonadotropin levels greater than 2.0 multiples of the median and 26 (10.2%) had alpha-fetoprotein levels greater than 2.5 multiples of the median. Of 11 pregnancies with fetal aneuploidy, 6 (54.5%) had human chorionic gonadotropin levels greater than 2.0 multiples of the median. It is concluded that in human chorionic gonadotropin screening programs for fetal Down syndrome, weight and race adjustments are necessary for accurate risk assessment.

Aneuploidy↗
PubMed1991

Intestinal vitamin D-dependent calbindin-D9k and alkaline phosphatase in spontaneously hypertensive rats.

Intestinal Ca2+ malabsorption has been described in spontaneously hypertensive rats (SHRs), but the molecular basis for this defect is unknown. In this study, we measured intestinal alkaline phosphatase and vitamin D-dependent Ca(2+)-binding protein (calbindin-D9k), two proteins implicated in the active pathway of intestinal Ca2+ absorption. Both proteins were measured in the small intestines of SHRs and their normotensive controls, Wistar-Kyoto rats, before, during, and after development of hypertension (4, 9, 14, 18, and 28 wk of age). At all ages, alkaline phosphatase activity in duodenum (0-6 cm) was decreased by 30-57% (P less than 0.001) and by 47-75% in the 2nd intestinal segment (6-12 cm) (P less than 0.001-0.05). Calbindin-D9k was decreased similarly. The decreases of calbindin were statistically significant (P less than 0.001-0.05) in the duodena at 4, 14, 18, and 28 wk (9-30% decreases) and in the 2nd segment at 4, 14, and 18 wk (38-69% decreases; P less than 0.001-0.005). Decreased calbindin in SHRs was documented in animals from two suppliers. The deficiencies of calbindin-D9k and alkaline phosphatase could not be attributed to malnutrition or to a generalized brush-border defect as indicated by body weights and the intestinal marker enzyme sucrase. Although calbindin-D9k was decreased in young SHRs, the serum 1,25-dihydroxycholecalciferol [1,25(OH)2D3] was increased by 59 and 129% in 4- and 9-wk-old SHRs (P less than 0.001), respectively; by contrast, serum 1,25(OH)2D3 was unchanged or decreased in older SHRs.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1991

DA1 dopamine receptors in renal cortical collecting duct.

Renal dopamine DA1 receptors are linked to the regulation of sodium transport. We have previously reported the presence of DA1 receptors in the proximal convoluted tubule (PCT) but not in the distal convoluted tubule. However, the DA1 receptor in the collecting duct, the final determinant of electrolyte transport, has not been studied. DA1 receptors were studied in the microdissected cortical collecting duct (CCD) of rats by autoradiography with use of the selective DA1 radioligand 125I-Sch 23982 and by measurement of adenylate cyclase (AC) activity. Specific binding of 125I-Sch 23982 to CCD was saturable with radioligand concentration. The dissociation constant (Kd) was 0.46 +/- 0.08 nM (n = 5), and the maximum receptor density (Bmax) was 1.41 +/- 0.43 fmol/mg protein (n = 5). The DA1 antagonist Sch 23390 was more effective than the DA1 agonist fenoldopam in competing for specific 125I-Sch 23982 binding. Fenoldopam stimulated AC activity in CCD in a concentration-dependent (10(-9)-10(-6) M) manner. The ability of fenoldopam to stimulate AC activity was similar in CCD and PCT even though DA1 receptor density was 1,000 times greater in the CCD than in the PCT. In additional studies, fenoldopam stimulation of AC activity did not influence vasopressin-stimulated AC activity. We conclude that the DA1 receptor in rat CCD is tightly coupled to AC stimulation and that there is no interaction between DA1 agonist-stimulated and vasopressin-stimulated AC activity in the CCD.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1991

Laboratory systems integration: robotics and automation.

Robotic technology is going to have a profound impact on the clinical laboratory of the future. Faced with increased pressure to reduce health care spending yet increase services to patients, many laboratories are looking for alternatives to the inflexible or "fixed" automation found in many clinical analyzers. Robots are being examined by many clinical pathologists as an attractive technology which can adapt to the constant changes in laboratory testing. Already, laboratory designs are being altered to accommodate robotics and automated specimen processors. However, the use of robotics and computer intelligence in the clinical laboratory is still in its infancy. Successful examples of robotic automation exist in several laboratories. Investigators have used robots to automate endocrine testing, high performance liquid chromatography, and specimen transportation. Large commercial laboratories are investigating the use of specimen processors which combine the use of fixed automation and robotics. Robotics have also reduced the exposure of medical technologists to specimens infected with viral pathogens. The successful examples of clinical robotics applications were a result of the cooperation of clinical chemists, engineers, and medical technologists. At the University of Virginia we have designed and implemented a robotic critical care laboratory. Initial clinical experience suggests that robotic performance is reliable, however, staff acceptance and utilization requires continuing education. We are also developing a robotic cyclosporine which promises to greatly reduce the labor costs of this analysis. The future will bring lab wide automation that will fully integrate computer artificial intelligence and robotics. Specimens will be transported by mobile robots. Specimen processing, aliquotting, and scheduling will be automated.(ABSTRACT TRUNCATED AT 250 WORDS)

Artificial Intelligence↗
PubMed1990

Molecular biology of adrenergic and dopamine receptors and the study of developmental nephrology.

Neurotransmitters convey specific messages by binding to receptors on the cell membrane surface. Receptors are linked to membrane-bound, signal-transducing proteins which act as intermediaries in the generation of second messengers that elicit biological responses. Cell surface receptors could be grouped into families that utilize common systems for their signal transmission. These classes include the growth factor receptors, the transporter receptors which internalize their ligands, ion channels, and G-protein-coupled receptors. In the past few years, the cDNAs and/or genes of a number of G-protein-coupled receptors have been cloned. Structural analysis of the G-protein-coupled receptors, as well as the other classes of receptor, shows that those receptors which use a common signaling pathway have similar topographies and share significant sequence homology. Adrenergic and dopamine receptors are examples of receptors coupled to G proteins. This review outlines some strategies in the study of adrenergic and dopamine receptors using molecular biology techniques and how they relate to investigations in developmental nephrology.

GTP-Binding Proteins↗
PubMed1990

Comparison of the central and renal dopamine-1 receptor.

The dopamine D-1 receptor from striatal membranes was compared with the dopamine DA-1 receptor from renal proximal tubules. The dopamine-1 receptors were solubilized with 1% sodium cholate and phospholipids after pretreatment with the dopamine-1 agonist, SKF R-38393. The soluble receptors were reconstituted into phospholipid vesicles after removal of sodium cholate. The receptors were studied by radioligand binding using the dopamine-1 antagonist [125I]-SCH 23982. The reconstituted dopamine D-1 and DA-1 receptor densities were similar. However, the affinity of the solubilized D-1 receptor was 17-fold greater than the solubilized DA-1 receptor. The affinity of membrane bound D-1 receptor to the radioligand was also greater than that noted for membrane bound DA-1 receptor. The mechanism for this difference remains to be determined.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1990

Dopamine-1 and dopamine-2 mechanisms in the control of renal function.

Dopamine (DA), a catecholamine produced in the kidney, is a renal vasodilator and natriuretic substance, but its action at dopamine-1 (DA-1), dopamine-2 (DA-2) and alpha- and beta-adrenergic receptors limits its effectiveness as a heuristic tool and pharmacologic agent. We have studied the effects of highly selective DA-1 and DA-2 receptor agonists and antagonists in normal human subjects and experimental animals to determine the precise physiological role of renal dopamine at DA-1 and DA-2 receptors within the kidney. We studied fenoldopam, a selective DA-1 agonist, in normal human subjects in metabolic balances at high (300 mEq/day) and low (10 mEq/day) sodium (Na) intake. Selective DA-1 receptor stimulation during high Na intake resulted in renal vasodilation, natriuresis and diuresis in a sustained manner for 3 hours. The natriuresis was mediated by a reduction in Na reabsorption at both proximal and distal tubular sites. In contrast, during low Na intake, DA-1 receptor stimulation did not engender a natriuretic or diuretic response. Thus, sodium depletion may inhibit the function of renal tubular cells in response to DA-1 stimulation. DA-1 receptors are present in the medial layer of the renal vasculature, proximal tubule and cortical collecting duct; DA-2 receptors are localized to the glomerulus, the renal nerves surrounding renal blood vessels and possibly the renal vascular endothelium. We have performed studies in conscious dogs with indwelling renal arterial catheters to identify the physiological role of renal DA to DA-1 and DA-2 receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1990

A renal dopamine-1 receptor defect in two genetic models of hypertension.

Dopamine (DA), via DA-1 receptors, regulates Na+ transport in the kidneys. Dopamine is synthesized from L-DOPA in the proximal tubule and presumably secreted as an autocrine/paracrine substance to stimulate DA-1 receptors localized on proximal tubular cells. We have previously reported the presence of DA-1 receptors in renal cortical homogenates and on the isolated proximal tubule of the rat and rabbit, consistent with the dopamine autocrine/paracrine model. We have localized DA-1 receptors in the proximal straight tubule of the rabbit, and in the cortical collecting duct of the rabbit and rat, but not in the distal collecting tubule or the cortical thick ascending loop of Henle. The presence of functional DA-1 receptors has been substantiated by the coexistence of DA-1 agonist-stimulated adenylate cyclase activity in the same nephron segments in which DA-r receptors have been found. Increased concentrations of intrarenal dopamine induced by dopamine-beta-hydroxylase inhibition with SKF-102698 caused a down regulation of proximal tubular DA-1 receptors and almost complete ablation of DA-1 agonist stimulated adenylate cyclase activity. Thus, dopamine may play a role in the regulation of DA-1 receptors and their linkage with adenylate cyclase. Since alterations in the renal dopaminergic system have been measured in some forms of experimental hypertension, we studied DA-1 receptors and their coupling to adenylate cyclase in the spontaneously hypertensive rat (SHR).(ABSTRACT TRUNCATED AT 250 WORDS)

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1990

Physiological modulation of renal function by the renal dopaminergic system.

1. The renal dopaminergic system is a potentially important regulator of sodium homeostasis and kidney function. 2. We have presented evidence that dopamine acts as a paracrine substance at DA-1 and DA-2 receptors in the physiological control of renal function. 3. Much more information is required regarding basic cellular mechanisms and the functional regulation of the system so that the role of renal dopamine can be placed clearly in context with other established hormonal regulatory systems.

Animals↗