Biomedical subjects

M J Peach

Publications and source records attributed to M J Peach.

At least 37 records · Page 2Linked to original sources

PubMed1991

Role of intrarenal angiotensin II and alpha-adrenoceptors in renal vasoconstriction with acute hypoxemia and hypercapnic acidosis in conscious dogs.

To evaluate our previous observation of renal vasoconstriction during combined acute hypoxemia and hypercapnic acidosis preceded by acute hypoxemia, we studied 13 conscious mongrel uninephrectomized dogs with chronic renal catheters and controlled sodium intake (80 meq/day for 4 days). Five dogs were studied during combined acute hypoxemia (PaO2, 37 +/- 1 mm Hg) and hypercapnic acidosis (PaCO2, 59 +/- 1 mm Hg; pH 7.20 +/- 0.01). Each dog was studied during infusion of 1) the intrarenal vehicle (n = 5), 2) the intrarenal alpha 1-antagonist prazosin (0.2 micrograms.kg-1.min-1, n = 5), 3) intrarenal [Sar1,Ala8]angiotensin II (70 ng.kg-1.min-1, n = 5), and 4) intrarenal prazosin and [Sar1,Ala8]angiotensin II (n = 4). Immediate induction of combined hypoxemia and hypercapnic acidosis after control measurements during intrarenal vehicle infusion resulted in a decrease in effective renal plasma flow and glomerular filtration rate, increase in renal vascular resistance, and decrease in filtered sodium load in the first 20 minutes of the blood gas derangement. Intrarenal administration of [Sar1,Ala8]angiotensin II failed to reverse the effects of the combined blood gas derangement on renal function. In contrast, intrarenal prazosin administration either alone or in combination with [Sar1,Ala8]angiotensin II abrogated the increase in renal vascular resistance, decrease in glomerular filtration rate, and fall in filtered sodium load. These studies identify a major role for alpha 1-adrenoceptors in the renal vasoconstriction during combined hypoxemia and hypercapnic acidosis.

Acidosis↗

PubMed1991

Renal nerves modulate renin gene expression in the developing rat kidney with ureteral obstruction.

Chronic unilateral ureteral obstruction (UUO) in newborn rats activates renin gene expression in the obstructed kidney, and increases renin distribution along afferent glomerular arterioles in both kidneys. To investigate the role of the renal nerves in this response, 2-d-old Sprague-Dawley rats were subjected to UUO or sham operation. Chemical sympathectomy was performed by injection of guanethidine, whereas, control groups received saline vehicle. At 4-5 wk, renal renin distribution was determined by immunocytochemistry, and renin mRNA levels were determined by Northern blot hybridization. Compared to the saline-treated rats with UUO, renin remained localized to the juxtaglomerular region in both kidneys of rats with UUO receiving guanethidine (P less than 0.05). Moreover, renin mRNA levels were eightfold lower in obstructed kidneys of rats receiving guanethidine than in those receiving saline. Additional groups of rats with UUO were subjected to unilateral mechanical renal denervation: renin gene expression in the obstructed kidney was suppressed by ipsilateral but not by contralateral renal denervation. These findings indicate that either chemical or mechanical denervation suppressed the increase in renin gene expression of the neonatal kidney with ipsilateral UUO. We conclude that the renal sympathetic nerves modulate renin gene expression in the developing kidney with chronic UUO.

Animals↗

PubMed1990

Expression of renin and its mRNA in the adult rat kidney with chronic ureteral obstruction.

Angiotensin II has been implicated in mediating renal vasoconstriction resulting from chronic unilateral ureteral obstruction (UUO) in both mature and developing animals. We have previously shown that chronic neonatal UUO results in increased distribution of renin and its mRNA in the obstructed kidney, as well as of immunoreactive renin in the intact opposite kidney. The present study was designed to evaluate the effects of 24 hours versus 4 weeks of UUO on the distribution of renin mRNA and its protein in the adult rat kidney. Renin was detected by immunocytochemistry using a polyclonal anti-rat renin antibody. Renin mRNA was localized by in situ hybridization to an oligonucleotide complementary to renin mRNA. UUO of 24 hours' or 4 weeks' duration did not alter the distribution of renin and its mRNA in the obstructed kidneys as compared with sham-operated kidneys, although kidneys obstructed for 4 weeks had a significant increase in the percent of renin-containing juxtaglomerular apparatuses (JCA) when compared with the intact opposite kidneys (P less than 0.05). Compensatory hypertrophy was not present in the intact opposite kidneys after 24 hours of UUO and distribution of renin gene expression was not altered at that time. However, 4 weeks following contralateral UUO, the intact kidneys were hypertrophied and showed a decrease in renin gene expression relative to the obstructed and sham-operated kidneys. We conclude that unlike UUO during early development, chronic UUO in the mature animal does not activate renin gene expression nor alter renin distribution in the obstructed kidneys. Renin gene expression is suppressed in the hypertrophied kidney with prolonged contralateral UUO.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

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

The angiotensin II receptor and the actions of angiotensin II.

Angiotensin II (Ang II) is a potent effector peptide of the renin-angiotensin system that exerts a wide variety of physiological actions on the cardiovascular, renal, endocrine, and central and peripheral nervous systems. Angiotensin exerts its actions by binding to specific receptors in the plasma membrane of various tissues. Structure-activity relationship studies and competition-binding experiments have identified a potency series of angiotensin analogues. Such studies have demonstrated that target organs display different preferences for Ang II and homologues such as Ang III and des-[Phe8] angiotensin II. Similarly, agents that normally are considered to be pure receptor antagonists for a given response (tissue) are full agonists in other tissues. Indirect evidence obtained from the above studies have led to the speculation that there are multiple angiotensin receptor subtypes among various tissues as well as within single cell types. Multiple mechanisms of signal transduction have been demonstrated for angiotensin. For example, depending on the effector organ, angiotensin stimulates phosphoinositide turnover and release of internal calcium, modulates voltage-dependent calcium channels, directly activates calcium channels, and inhibits adenylate cyclase activity. Recently, the identification of selective, high-affinity peptide and nonpeptide antagonists has resulted in further characterization of angiotensin receptors into distinct subtypes. In addition, dithiothreitol, an agent that reduces disulfide bridges, has been a useful tool in the characterization of angiotensin receptors as the subtypes apparently are not affected equally by this agent. However, further work needs to be performed to characterize angiotensin receptors with respect to heterogeneity, structure, transducing mechanisms, and physiological function.

Angiotensin II↗

PubMed1990

Microcirculatory responses to exogenous endothelial cell-derived relaxing factor.

Endothelium-derived relaxing factor (EDRF) plays an important role in the vasodilatory responses of large blood vessels. However, such a role has yet to be conclusively shown for the microvasculature. In this study we tested the sensitivity of arterioles in the cheek pouch of pentobarbital-anesthetized hamsters to the EDRF-dependent agonists bradykinin and A23187, as well as to exogenous EDRF from cultured bovine aortic endothelial cells. The pouch superfusion fluid was arranged to first pass through a column containing endothelial cells and then on to the tissue. Bradykinin (10-30 nM) or A23187 (0.3 microM) was introduced either upstream or downstream to the endothelial cells, and the resultant responses were measured with video microscopy. Bradykinin and A23187 both caused a dose-dependent release of a microvessel dilator from cultured endothelial cells. We take this dilator to be EDRF based on the characteristics of the responses to the stimuli. Indomethacin (7.7 microM) was present in the superfusate to eliminate the production of cyclooxygenase products from the endothelial cells, and the magnitude of the response was diminished if the superfusate was first passed through a 3-min delay coil before arrival at the pouch. The arterioles dilated to the direct application of bradykinin in a dose-dependent fashion. They did not respond however to the direct application of A23187. These studies demonstrate that arteriolar smooth muscle is able to respond to exogenous EDRF and support the premise that EDRF may play an active role in the regulation of blood flow in the microcirculation.

Animals↗

PubMed1990

Hematocrit modulates response of ANP to volume expansion in immature rats.

Compared with the adult, the immature kidney responds to acute volume expansion (VE) with reduced diuresis and natriuresis. The underlying mechanisms are poorly understood. Diuresis and natriuresis are blunted despite a lower hematocrit (Hct) in the immature rat, which should enhance the response to VE. The present study was designed to evaluate the role of atrial natriuretic peptide (ANP), its second messenger, guanosine 3',5'-cyclic monophosphate (cGMP), and Hct in the regulation of the renal response to acute saline VE (3% body wt) during postnatal development in rats. Anesthetized Sprague-Dawley rats were studied in the preweaning (group P) or postweaning period (group W). In additional groups, Hct was increased in preweaned rats (group PI) and decreased in postweaned rats (group WD) by isovolemic exchange transfusion before VE. Although control plasma ANP concentration [( ANP]) was not different among the four groups, the increased diuretic and natriuretic response to VE in group W was associated with a greater increase in [ANP] and urinary cGMP excretion (UcGMPV) than in group P (P less than 0.05). Experimental increases in Hct in group PI resulted in greater [ANP] after VE but a reduction in UcGMPV and diuresis (P less than 0.05), whereas decrease in Hct in group WD increased UcGMPV without a significant effect on [ANP], diuresis, or natriuresis. We conclude that ANP may contribute to the increasing renal response to VE with maturation and that Hct modulates ANP release, UcGMPV, and the renal response to VE in postnatal maturation.

Aging↗

PubMed1990

Combined intrarenal blockade of the renin-angiotensin system in the conscious dog.

We produced maximal or near-maximal acute intrarenal blockade of the renin-angiotensin system (RAS) by combining inhibitors. Intrarenal infusion of the renin inhibitor, ACRIP, the converting enzyme inhibitor, teprotide, and saralasin were administered individually or combined in random order. The inhibitors were infused for 20 min in doses that did not produce systemic effects in uninephrectomized conscious dogs in sodium balance at 10 meq/day. Significant increases in urine flow rate (UV; F = 97, P less than 0.0001), urinary sodium excretion (UNaV; F = 220, P less than 0.0001), glomerular filtration rate (GFR; F = 64, P less than 0.0001), and renal plasma flow (RPF; F = 108, P less than 0.0001) were observed with each blocker, whether alone or in combination except that ACRIP alone did not alter GFR or RPF. The increase in renal function was related to the number of blockers (3 greater than 2 greater than 1). With the three blockers combined UV increased approximately sixfold (from 0.5 +/- 0.06 to 2.9 +/- 0.03 ml/min), UNaV approximately 10-fold (from 3 +/- 0.4 to 34 +/- 2.8 mueq/min), GFR from 31 +/- 2 to 49 +/- 2 ml/min, RPF from 59 +/- 1 to 120 +/- 4 ml/min, and fractional excretion of sodium from 0.06 +/- 0.01 to 0.5 +/- 0.4% (all P less than 0.001). These changes did not occur where the inhibitors were infused systemically and the changes during intrarenal blocker administration were blocked completely with co-administration of angiotensin II intrarenally. The intrarenal RAS is a potent physiological regulator of renal function.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗

PubMed1990

Identification of individual renocortical cells that secrete renin.

Successful application of the reverse hemolytic plaque assay was developed to identify individual renocortical cells that secrete renin directly. The plaque assay was validated by a number of established criteria. Using this technique, we demonstrate an increase in renin secretion with beta-adrenergic stimulation and an inhibition of renin secretion with extracellular calcium in groups of renin-secreting cells. Transmission electron microscopy of the cell in the center of a hemolytic plaque demonstrated a modified vascular smooth muscle cell with densely packed secretory granules. Electron microscopy immunocytochemistry demonstrated the presence of renin in the secretory granules, confirming the identity of the cell as a renal juxtaglomerular cell. The technology developed here has allowed the precise identification and study of the individual renin-secreting juxtaglomerular cell.

Animals↗

PubMed1990

In situ localization of renin and its mRNA in neonatal ureteral obstruction.

Angiotensin II is an important mediator of renal vasoconstriction resulting from chronic unilateral ureteral obstruction (UUO). Distribution of renin mRNA and immunoreactive renin (IR) was examined in kidneys of 1-mo-old Sprague-Dawley rats subjected to either sham operation (n = 21), left complete UUO (n = 21), or right uninephrectomy (UNX, n = 16) at 2 days of age. There were no differences among the three groups in mean arterial pressure or plasma renin activity. Unlike sham kidneys, in which IR was detected in less than 55% of juxtaglomerular apparatuses (JGA) and was confined to a juxtaglomerular location, IR in both kidneys of animals with UUO appeared in greater than 75% of JGA and extended along most of the length of the afferent arteriole (P less than 0.01). In contrast, IR in kidneys of UNX rats was localized to the JGA as in sham-operated animals. Compared with sham-operated kidneys, renal renin content was increased in the obstructed kidneys (P less than 0.01) but decreased in the intact opposite kidneys of UUO rats and in the remaining kidneys of UNX rats (P less than 0.05). Renin mRNA, detected by in situ hybridization histochemistry, was localized to the JGA in kidneys of all groups. However, the fraction of JGA containing detectable renin mRNA was higher in obstructed kidneys than in intact opposite, UNX, or sham kidneys (P less than 0.05). In conclusion, UUO alters intrarenal renin independent of the systemic renin-angiotensin system. The greater distribution of IR, increased renin content, and renin gene expression of kidneys with ipsilateral UUO are consistent with a role for renin-angiotensin in mediating the vasoconstriction resulting from UUO.

Animals↗

PubMed1990

Recruitment of renin gene-expressing cells in adult rat kidneys.

To define whether angiotensin I-converting enzyme (ACE) inhibition affects the distribution of renin gene-expressing cells within the kidney, a control group of adult male Wistar-Kyoto rats (C, n = 7) was compared with a group of rats treated with enalapril (E, n = 6) for 5 days. Renin mRNA distribution was assessed using in situ hybridization to a 35S-labeled 28 mer oligonucleotide complementary to rat renin mRNA. Whereas in control rats renin mRNA was confined to a juxtaglomerular location, in enalapril-treated rats, renin mRNA extended proximally along the length of the afferent arteriole. The percent of visible afferent arteriolar length containing renin mRNA was higher in enalapril-treated (71.7 +/- 2.8%) than in control (49.6 +/- 2.1%) rats (P less than 0.0001). These findings were accompanied by an increase in the percent of juxtaglomerular apparatuses (JGAs) containing renin mRNA (71 +/- 2.2 vs. 49 +/- 2.9%; E vs. C, P less than 0.0001). Also, the intensity of the JGA hybridization signals was higher in enalapril-treated (757 +/- 59 grains/JGA) than in control (167 +/- 11 grains/JGA) rats (P less than 0.00001). We conclude that the increased kidney renin gene expression elicited by ACE inhibition is the result of an increase in renin mRNA content per JGA, an increase in the number of JGAs expressing the renin gene, and a recruitment of renin gene-expressing cells along the afferent arteriole.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1990

Evidence that dopamine-2 mechanisms control renal function.

Dopamine is synthesized by the kidney, and dopamine-2 (DA2) receptors are present in the renal glomerulus. However, no role for DA2 receptors in the kidney has been defined. We investigated the possible role of DA2 receptors in control of renal function by intrarenal infusion of a highly specific DA2 antagonist YM-09151 (YM), in conscious uninephrectomized dogs (n = 5) in metabolic balance at Na intake 40 meq/day. YM infused at 0.01 pmol.kg-1.min-1 did not cause any changes in urinary flow rate or Na excretion. Administration of YM (infusions from 0.1 to 10.0 pmol.kg-1.min-1) caused a significant dose-dependent diuresis (F = 20.3; P less than 0.001) and natriuresis (F = 35.2; P less than 0.0001) and an increase in glomerular filtration rate (F = 45.4; P less than 0.0001), renal plasma flow (F = 209.3; P less than 0.0001), and filtration fraction (F = 11.2; P less than 0.0001). No significant changes in plasma renin activity, plasma aldosterone concentration, or mean arterial blood pressure occurred with any of the doses of YM infused into the renal artery. Coinfusion of LY-171555, a specific DA2 agonist, at a dose that itself did not affect renal function, completely abrogated the renal hemodynamic and excretory changes induced by YM. The data suggest that dopamine produced intrarenally may act at renal vascular and/or glomerular DA2 receptors to control renal function.

Animals↗

PubMed1990

Angiotensin II regulates renin gene expression.

We investigated the effect of angiotensin II (ANG II) and enalapril on accumulation of renin messenger RNA (mRNA) and on renal renin distribution (immunohistochemical analysis). Adult Wistar-Kyoto rats received enalapril (0.2 mg/ml) in distilled drinking water for 8 or 12 days. On day 5 of enalapril treatment, an osmotic minipump was implanted in the peritoneum that caused sustained release of ANG II (200 ng.kg-1.min-1) or vehicle (bovine serum albumin) for 3 or 7 days. Control rats received water for 8 or 12 days and osmotic minipump implantation (containing vehicle solution) on the 5th day. Renin mRNA was identified by hybridization with a 32P-labeled full-length complementary DNA and was detected by autoradiography. Enalapril treatment increased renal renin mRNA specific activity (renin mRNA/total RNA). Subsequent infusion of angiotensin II for 3 or 7 days decreased renal renin mRNA specific activity. In addition, renin immunostaining increased along the afferent arteriole after enalapril treatment; however, enalapril-induced spread of renin immunostaining was not inhibited by ANG II. Thus ANG II attenuates the accumulation of renin mRNA stimulated by enalapril treatment without alteration of renal renin distribution. The lack of effect of ANG II on renal renin distribution may be due to the length of turnover time for stored protein. These findings suggest the shortloop negative feedback of ANG II on renin reflects inhibition of renin synthesis by ANG II. Therefore, we propose that ANG II exerts a direct inhibitory effect on renin by regulation of renin gene expression in renal vasculature.

Aldosterone↗

PubMed1990

Regulation of cytosolic calcium by angiotensins in vascular smooth muscle.

The carboxy terminal homologue of angiotensin II (Ang II), Ang-(3-8) or hexapeptide, was used as a model peptide to examine the types of receptor mechanisms involved in calcium mobilization in cultured vascular smooth muscle cells. Hexapeptide did not produce tachyphylaxis but did produce a sustained increase in intracellular calcium. Differences in the increase in intracellular calcium [( Ca2+]i) and the pattern of inositol phosphate production indicate that Ang-(3-8) and maximal concentrations of Ang II mobilize calcium through different mechanisms. The calcium-mobilizing mechanisms that predominate appear to depend on the concentration of angiotensin. Concentrations of Ang II greater than 10(-8) M produce sharp calcium transients in which the [Ca2+]i returns close to baseline within 1 minute after stimulation, but concentrations of Ang II equal to or less than 3 x 10(-9) M result in a plateau increase in calcium. Pretreatment with Bordetella pertussis toxin does not abolish either the calcium transient induced by Ang II or the plateau phase induced by Ang-(3-8), indicating that the GTP-transducing protein that couples the receptor to phospholipase C or, possibly, a receptor-operated calcium channel is not Bordetella pertussis toxin sensitive.

Angiotensins↗

PubMed1990

Changes in angiotensinogen messenger RNA in differentiating 3T3-F442A adipocytes.

Angiotensinogen messenger RNA (mRNA) has been identified in both brown and white adipose tissue. Recently we have shown that when 3T3-L1 cells were treated with isobutylmethylxanthine (IBMX) to accelerate differentiation, angiotensinogen mRNA increased markedly in adipocytes as compared with preadipocytes. To determine if a correlation existed between the regulatory events associated with the differentiation process, we compared the change in angiotensinogen mRNA in spontaneously differentiating 3T3-F442A cells with two established parameters of differentiation in adipocyte cell lines. Differentiation was assessed by visual examination of cells for lipid droplets, fluorescent staining of the F-actin fibers, and increases in glycerol phosphate dehydrogenase mRNA. F-actin fibers were highly structured in preadipocytes, becoming disassembled and very disorganized as cells differentiated into adipocytes. The quantity of angiotensinogen mRNA increased as the number of lipid-containing cells increased within a culture. Glycerol phosphate dehydrogenase mRNA accumulated in differentiated adipocytes to about the same extent as angiotensinogen mRNA. Thus, increases in angiotensinogen mRNA were associated with the morphological and biochemical changes that occur during the phenotypic modulation of 3T3-F442A cells.

Actins↗

PubMed1990

Selective peripheral dopamine-1 receptor stimulation. Differential responses to sodium loading and depletion in humans.

Dopamine-1 (DA1) receptors in the renal tubules may be involved in the regulation of sodium homeostasis. To test this hypothesis, fenoldopam, a selective DA1 agonist, was infused at 0.05 microgram/kg/min i.v. in 16 normal male subjects in metabolic balance at 300 or 10 meq sodium. Renal function studies were performed by standard p-aminohippurate, inulin, and lithium clearances for three periods: 1) precontrol (2 hours), 2) experimental (3 hours), and 3) postcontrol (2 hours). DA1 receptor stimulation in sodium-loaded individuals increased the following parameters during the experimental period: urine flow rate, from 12.5 +/- 0.4 to 15.5 +/- 0.5 ml/min (p less than 0.05); urinary sodium excretion, from 309 +/- 12 to 489 +/- 18 mu eq/min (p less than 0.001); renal plasma flow, from 631 +/- 19 to 717 +/- 21 ml/min (p less than 0.005); fractional sodium excretion, from 2.2 +/- 0.1% to 3.4 +/- 0.1% (p less than 0.001); fractional lithium excretion, from 26.2 +/- 0.7% to 32.1 +/- 0.8% (p less than 0.005); and distal sodium load, from 10.7 +/- 0.4 to 13.8 +/- 0.5 ml/min (p less than 0.05). The increase in fractional sodium excretion was greater than that of fractional lithium excretion (p less than 0.0001). Distal sodium reabsorption decreased from 78.3 +/- 0.8% to 73.2 +/- 1.1% but the change was not statistically significant. In contrast, sodium-depleted subjects exhibited no significant changes except in renal plasma flow, which rose from 550 +/- 13 to 625 +/- 17 ml/min (p less than 0.0001). Glomerular filtration rate remained unchanged through the entire study. These results indicate that diuretic and natriuretic responses are mediated by DA1 receptors at both proximal and distal tubular sites. Attenuation of the DA1 natriuretic response during sodium depletion suggests a direct inhibition of cellular DA1 mechanisms in the renal tubule or recruitment of nondopaminergic compensatory homeostatic mechanisms within the kidney.

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

PubMed1990

NG-monomethyl L-arginine inhibits endothelium-derived relaxing factor-stimulated cyclic GMP accumulation in cocultures of endothelial and vascular smooth muscle cells by an action specific to the endothelial cell.

The effect of NG-monomethyl L-arginine (LNMMA), an analogue of L-arginine (a proposed precursor of endothelium-derived relaxing factor [EDRF]), on EDRF release from bovine pulmonary artery endothelial cells was investigated using endothelial cell-vascular smooth muscle cocultures and a superfused column containing endothelial cells grown on microcarrier beads. Cocultures were stimulated with control buffer, ATP, bradykinin, melittin, A23187, or nitroprusside in the presence and absence of varying concentrations of LNMMA (30-300 microM). LNMMA caused significant, concentration-dependent decreases in cyclic GMP accumulation in response to the endothelium-dependent dilators bradykinin, ATP, melittin, and A23187 but had no effect on control or nitroprusside-stimulated cocultures. The inhibitory effect of LNMMA on cyclic GMP accumulation was partially reversed by treatment with L-arginine, but was unaffected by D-arginine. To determine the specific site of action of LNMMA, endothelial cells on microcarrier beads were placed in a column and superfused with buffer. The effluent from the column was collected in 30-second (1.5-ml) fractions into 2-cm2 monolayer wells of vascular smooth muscle cells before and after addition of agonists (bradykinin, A23187) to the column inflow. The cyclic GMP content of each well of smooth muscle cells was determined as an index of EDRF activity. LNMMA superfused through the endothelial cell column inhibited cyclic GMP accumulation in vascular smooth muscle cells induced by bradykinin and A23187. LNMMA introduced into the effluent from the endothelial cell column had no effect on smooth muscle cyclic GMP levels. We conclude that LNMMA is an effective, specific inhibitor of EDRF production or release, and its action is specific to the endothelial cell.

Adenosine Triphosphate↗

PubMed1990

Renin release and gene expression in intact rat kidney microvessels and single cells.

To investigate whether newborn kidney microvessels and isolated single microvascular cells have the capacity to release renin and/or alter the expression of the renin gene in response to adenylate cyclase stimulation, newborn kidney microvessels were isolated and purified (95%) using an iron perfusion/enzymatic digestion technique. Incubation of microvessels with either vehicle (control; C) or 10(-5) M forskolin (F) in media resulted in an increase in microvessel cAMP (0.67 +/- 0.13 vs. 22 +/- 4.6 pmol/min per mg protein) (P less than 0.005) and renin released into the culture media (1,026 +/- 98 vs. 1,552 +/- 159 pg angiotensin I/h per mg protein) (P = 0.008) (C vs. F). Renin mRNA levels in the newborn kidney microvessels increased 1.6-fold with forskolin treatment. Renin release by isolated, single microvascular cells (with or without forskolin) was assessed using the reverse hemolytic plaque assay. Forskolin administration resulted in an increase in the number of renin-secreting cells without changes in the amount of renin secreted by individual cells. In conclusion, newborn kidney microvessels and isolated renin-releasing microvascular cells possess a functionally active adenylate cyclase whose short-term stimulation results in accumulation of cAMP, a significant increase in renin release, and an enhancement of renin gene expression. The increase in renin release is due to recruitment of microvascular cells secreting renin. Recruitment of hormone-secreting cells in response to stimuli may prove to be a mechanism of general biological importance shared by many endocrine cell types.

Animals↗