Biomedical subjects

J W Strandhoy

Publications and source records attributed to J W Strandhoy.

At least 19 recordsLinked to original sources

PubMed2001

Transendothelial permeability of chlorpyrifos in RBE4 monolayers is modulated by astrocyte-conditioned medium.

The immortalized rat brain endothelium 4 (RBE4) cell line preserves many features of the in vivo brain endothelium. It has been used as an in vitro model of the blood-brain barrier (BBB). Astrocyte-endothelial cell interactions are crucial for maintenance of BBB characteristics. The present study investigated morphological and permeability properties of the RBE4 cell line. Immunohistochemical studies showed positive staining in RBE4 cells for E-cadherin, a Ca(2+)-dependent cell-cell adhesion molecule. Western blot immunoassay showed that RBE4 cells consistently express E-cadherin and that its expression significantly increased (P<0.001) in the presence of astrocyte-conditioned medium (ACM). The transendothelial permeability of chlorpyrifos, an organophosphorus insecticide, was significantly decreased (P<0.001) when the RBE4 cells were grown in ACM compared with control medium. Additional studies were carried out to determine whether chlorpyrifos is a substrate for the multidrug resistance protein, P-glycoprotein (P-gp). No significant change in chlorpyrifos transendothelial permeability was noted in the presence of verapamil, a P-gp blocker. Thus, in this system, chlorpyrifos is not a substrate for P-gp. This work demonstrates that with additional refinements the RBE4 monolayers might serve as a useful in vitro model for the study of BBB permeability and modulation by astrocyte-derived soluble factors.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2000

Role of alpha-1 adrenoceptor subtypes mediating constriction of the rabbit ear thermoregulatory microvasculature.

An acute in vivo preparation of the microvasculature of the rabbit ear was used to evaluate the functional role of alpha1 (alpha1)-adrenoceptor subtypes in thermoregulatory microcirculation. The effect of alpha1-adrenoceptor subtype blockade on phenylephrine-induced vasoconstriction was assessed with the alpha1A, alpha1B, and alpha1D-adrenoceptor-selective antagonists 5-methyl-urapidil (10(-8) M), chloroethylclonidine (10(-5) M), and 8-[2-[4(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspirol[4.5]deca ne-7,9-dione dihydrochloride (BMY7378) (10(-6) M), respectively. The results demonstrated that pretreatment of the ear microvasculature with 5-methyl-urapidil or BMY7378 shifted the phenylephrine concentration-response curve rightward and significantly changed the log of the phenylephrine concentration, causing half-maximum stimulation (EC50) in arterioles (p < 0.05). BMY7378 shifted the phenylephrine concentration-response curve of the arteriovenous anastomoses about 100-fold rightward (p < 0.05). All three alpha1-adrenoceptor antagonists eliminated the vasoconstrictive effects of phenylephrine on venules. The results indicate that the ear microvasculature has a heterogenous distribution of alpha1-adrenoceptor subtypes. The alpha1A and alpha1D-adrenoceptor subtypes appear to have a greater influence on constrictive function in arterioles, whereas the alpha1D-adrenoceptor is the dominant constrictor of arteriovenous anastomoses. In general, the alpha1-adrenoceptor does not play a major vasoconstrictor role in venules. Chloroethylclonidine, an irreversible alpha1B-adrenoceptor antagonist, induced contractile responses in the ear microvasculature, probably due to its alpha2-adrenoceptor agonist effects. This study extended our understanding of the adrenergic receptor control mechanisms of a cutaneous thermoregulatory end organ characterized by two parallel perfusion circuits providing nutritional and thermoregulatory functions.

Animals↗

PubMed1999

Regulation of in vitro renin secretion by ANG II feedback manipulation in vivo in the ovine fetus.

The renin-angiotensin system is critically important to fetal cardiovascular function and organ development. The feedback regulation of renin secretion by ANG II develops early in gestation yet does not linearly progress from fetal life to adulthood. Renin secretion is elevated in late gestation compared with earlier or postnatal time periods, which suggests that some component of the negative feedback regulation of renin secretion is less sensitive in late gestation. We examined in fetal sheep the age-related consequence of chronic in vivo manipulation of ANG II on renal renin secretion measured in vitro. Immature (101-103 days of gestation) and mature (130-133 days of gestation) fetuses were treated for 72 h with enalaprilat, ANG II or vehicle. Content and basal and isoproterenol-stimulated secretion of prorenin (PR) and active renin (AR) from fetal kidney cortical slices were determined. Enalaprilat pretreatment in vivo increased renal renin content and basal and stimulated secretion of PR and AR in vitro even in immature animals. Immunohistochemical localization showed that enalaprilat treatment caused an age-related recruitment of renin-containing juxtaglomerular cells. Conversely, ANG II pretreatment decreased basal and stimulated PR and AR secretion from immature fetal kidneys, but only inhibited PR secretion from mature kidneys. It also caused an age-related decrease in the percentage of renin-containing juxtaglomerular cells. These results suggest that ANG II feedback modulates not only the synthesis and content of renin, but the sensitivity of the coupling between stimulus and secretion. A critical observation of our study is that the higher renal tissue concentrations of prorenin and active renin in late gestation may be a consequence of reduced sensitivity to ANG II feedback; this is consistent with the increased plasma concentrations of renin found in near-term mammals.

Angiotensin II↗

PubMed1998

Effect of cooling on cutaneous microvascular adrenoceptors in vivo in the rabbit ear.

Previous studies have suggested that moderate cooling increases the responsiveness of vascular alpha2-adrenoceptors. However, limited information is available documenting the influence of temperature changes on adrenoceptor responses in the microvasculature of thermoregulatory organs (e.g., the human digit and the rabbit ear) subjected to a wide range of temperatures. In the present study, the effect of local cooling (24 degrees C) on cutaneous microvascular adrenoceptors in the ear was observed in vivo in male New Zealand White rabbits (total: 66 ears). The rabbit ear was studied in a temperature-controlled tissue bath; the ear preparation was pretreated with terazosin (an alpha1-adrenoceptor antagonist) (10(-5) M) or a combination of terazosin (10(-5) M) and propranolol (a beta-adrenoceptor antagonist) (10(-6) M). The microvascular diameter responses of the ear to norepinephrine (10(-11)-10(-4) M) then were determined at 24 or 34 degrees C, respectively, to determine the influences of low temperature on adrenoceptor responses to norepinephrine stimulation. The results demonstrated that low concentrations of norepinephrine induced vasodilation in arterioles and arteriovenous anastomoses. This vasodilation was followed by vasoconstriction with an increased concentration of norepinephrine in animals with alpha1-adrenergic blockade at 34 degrees C. Moderate tissue cooling increased the microvascular maximal response of the rabbit ear to norepinephrine and abolished the vasodilatation induced by a low concentration of norepinephrine. There was no significant difference in the microvascular response to norepinephrine between the two temperature conditions after simultaneous blockade of alpha1-adrenoceptors and beta-adrenoceptors. Data from the present study indicate that moderate cooling does not enhance the responsiveness of alpha2-adrenoceptors to norepinephrine. In contrast, cooling reduced the beta-adrenergic activity of arterioles and arteriovenous anastomoses after norepinephrine stimulation.

Adrenergic alpha-Agonists↗

PubMed1996

Renal actions of angiotensin-(1-7): in vivo and in vitro studies.

In vivo studies were conducted in Na-replete anesthetized male Wistar rats with denervated kidneys. Intrarenal injections of angiotensin-(1-7) [ANG-(1-7) at > 1 nmol/kg produced a shallow dose-dependent decrease in renal blood flow that was mediated by the AT1-type ANG II receptor. A constant intrarenal infusion of ANG-(1-7) at 0.1 and 1 nmol.min-1.kg-1 had minimal effects on renal blood flow and blood pressure and resulted in an elevated urinary excretion of Na and water compared with the time-control saline-infused group. To determine whether ANG-(1-7) may have a direct action on tubular epithelium to inhibit Na reabsorption, we examined the effect of ANG-(1-7) on transport-dependent O2 consumption (Qo2) in fresh suspensions of rat proximal tubules in vitro. ANG-(1-7) inhibited Qo2 in a concentration-dependent fashion with a threshold concentration of approximately 100 pM. Stimulating Na-K-adenosinetriphosphatase (Na-K-ATPase) activity with nystatin caused a leftward shift of the inhibitory concentration-response curve to ANG-(1-7). The 22% inhibition of Qo2 by 1 pM ANG-(1-7) was abolished by pretreatment with 5 mM ouabain (Na-K-ATPase inhibitor), unaltered by pretreatment with 1 microM PD-123319 (AT2 receptor antagonist), partially attenuated by 1 microM losartan (AT1 receptor antagonist), and abolished by 1 microM [Sar1, Thr8]ANG II (nonselective ANG receptor antagonist). Together these findings indicate that ANG-(1-7) has biological activity in the kidney and, at nonvasoconstrictor doses, results in increased Na and water excretion in vivo. One site of action is the proximal tubule, where ANG-(1-7) can inhibit an ouabain-sensitive Na-K-ATPase exit step in cellular Na transport. This novel inhibitory action of ANG-(1-7) appears to be mediated by an AT1 receptor (minor component) and a non-AT1, non-AT2 ANG receptor (major component).

Angiotensin I↗

PubMed1996

Nitric oxide mediates the inhibitory action of platelet-activating factor on angiotensin II-induced renal vasoconstriction, in vivo.

The objective of our study was to determine the mechanism(s) involved in the inhibitory effect of platelet-activating factor on renal vascular reactivity, in vivo. Bolus injections of vasoconstrictor agonists were administered into the renal circulation of pentobarbital anesthetized male Wistar rats at a dose to cause a transient 45 to 50% decrease in renal blood flow. Intrarenal infusion of platelet-activating factor (PAF) at 2.5 ng/min/kg attenuated the vasoconstrictor response to angiotensin II by 66%, a significantly smaller reduction of 35% for norepinephrine-mediated vasoconstriction, 22% for vasopressin-mediated vasoconstriction and no alteration of KCl-mediated vasoconstriction. The preferential inhibitory effect of platelet-activating factor on angiotensin II-mediated renal vasoconstriction was mimicked by the intrarenal infusion of either 0.2 to 5 micrograms/min/kg methacholine (endothelium-dependent vasodilator) or 2 micrograms/min/kg sodium nitroprusside (nitric oxide donor). After inhibition of nitric oxide synthesis with NG-monomethyl-L-arginine, intrarenal infusion of PAF or methacholine reduced angiotensin II-mediated renal vasoconstriction significantly less than that observed in the absence of NG-monomethyl-L-arginine. Therefore, this study provides evidence that the shared ability of platelet-activating factor and methacholine to selectively reduce angiotensin II-mediated renal vasoconstriction involves endothelium-derived nitric oxide.

Angiotensin II↗

PubMed1993

Effects of phospholipids on renal function.

The effects of two classes of phospholipids (PL) on renal function have been studied. Bolus injections of 1 ng (10 pmol) of lysophosphatidylcholine (LPC) caused natriuresis and diuresis in rats. Natriuretic activity was eliminated by substituting unsaturated bonds in the 1-acyl group and by removing the choline group on the sn-3 position. Natriuretic activity was not affected by substitution of 1-alkyl for 1-acyl groups. In the dog, LPC was natriuretic when given as a bolus of 3.0 micrograms/kg or as a constant infusion at 5 ng/kg/min. To explore further the effect of alkyl PLs on renal function, a series of studies with platelet activating factor (PAF) was performed. PAF injected directly into the renal artery (IR) in bolus doses of 0.5-10 ng/kg caused renal vasodilation that was blocked by a specific PAF receptor antagonist. This effect was not due to release of vasodilatory eicosanoids, dopamine, or nitric oxide (NO). PAF given IR as a continuous infusion at 2.5 ng/kg/min attenuated the renal vasoconstrictor effects of angiotensin II and norepinephrine but not vasopressin. This effect to attenuate vasoconstriction was blocked by the NO inhibitor N-monomethyl-L-arginine. These studies using picomolar amounts of PL suggest a physiologic role for these compounds in control of renal function.

Angiotensin II↗

PubMed1992

Tetrodotoxin protects against acute ischemic renal failure in the rat.

Tetrodotoxin has been reported to cause prolonged systemic hypotension without resultant ischemic damage. We tested its ability to protect the kidney during 60 minutes of warm ischemia in uninephrectomized rats. Protection was observed when tetrodotoxin was given intravenously at two microgram./kg. and four microgram./kg. as assessed by serial plasma blood urea nitrogen and creatinine measurements over two weeks. Tetrodotoxin was protective when given immediately before or immediately after the ischemic period. The renal protection of tetrodotoxin was not due to its effects on renal nerves as renal denervation did not protect the kidney from the ischemic damage. The renal protective effects of four microgram. tetrodotoxin/kg. were similar to those of four mg. captopril/kg. but the combination of the two was paradoxically without effect. We tested whether tetrodotoxin and captopril chemically antagonized each other, but in the presence of tetrodotoxin, captopril was still a potent inhibitor of the conversion of angiotensin I to angiotensin II. These results indicate that tetrodotoxin could be useful in elucidating the sequence of events associated with ischemic-reperfusion renal injury and in identifying ways of preserving renal function during renal surgery.

Acute Kidney Injury↗

PubMed1991

Vasorelaxant effect of C16-PAF and C18-PAF on renal blood flow and systemic blood pressure in the anesthetized rat.

The renal vasoactive and systemic hypotensive effects of platelet activating factor (C16:0-PAF and C18:1-PAF) were examined in anesthetized male Wistar rats. Bolus injections of C16-PAF (0.5-25 ng/kg) and C18-PAF (2.5-200 ng/kg) into the arterial circulation of the kidney produced increases in renal blood flow (6-15%) before causing dose-dependent systemic hypotension (2-64 mmHg). The dose-response curves for renal blood flow and systemic blood pressure generated by intrarenal C18-PAF administration were approximately 7 fold to the right of the dose-response curves generated by C16-DPAF. Intrarenal injections of vehicle or the biologically inactive enantiomer C16-DPAF (25-200 ng/kg) did not affect renal blood flow or systemic blood pressure. These results suggest that C16:0-PAF is a more potent renal vasodilator and hypotensive lipid than C18:1-PAF.

Anesthesia↗

PubMed1991

Selective inhibition of vasoconstrictor responses by platelet-activating factor in rat kidney.

We examined the effect of platelet-activating factor (PAF) on renal vascular reactivity in the pentobarbital sodium-anesthetized male Wistar rat. Intrarenal infusion of C16-PAF at hypotensive (2.5 ng.min-1.kg-1) or nonhypotensive (0.5 ng.min-1.kg-1) doses caused renal vasodilation and dose dependently antagonized the renal vasoconstrictor responses of intrarenal boluses of angiotensin II (ANG II) greater than norepinephrine (NE) greater than vasopressin (AVP). PAF infusion at the high dose did not alter non-receptor-mediated renal vasoconstriction induced by intrarenal KCl injection. The inhibitory effect of PAF on agonist-induced renal vasoconstriction was accentuated by eicosanoid synthesis inhibition (indomethacin or dexamethasone), unaffected by dopamine-receptor blockade (haloperidol) but was totally abolished by PAF receptor antagonism (L-659,989). In contrast, intrarenal infusion of a calcium channel antagonist (nimodipine) or an intracellular calcium channel antagonist (TMB-8) equally inhibited the renal vasoconstrictor responses of ANG II, NE, and AVP. Thus PAF can cause renal vasodilation in the rat kidney and dose-dependently antagonizes the renal vasoconstrictor responses of ANG II greater than NE greater than AVP. The inhibitory effect of PAF on renal vasoconstrictor responses is mediated by PAF receptors and does not appear to be due to a nonspecific membrane effect, reduction in calcium mobilization, or the release of vasodilatory eicosanoids or dopamine.

Animals↗

PubMed1990

Dual interactions between alpha 2-adrenoceptor agonists and the proximal Na(+)-H+ exchanger.

In the kidney, the proximal nephron is a major site for Na+ reabsorption and H+ secretion. An electroneutral exchanger mediates the uptake of luminal Na+ with the secretion of cellular H+. In these studies, alpha-adrenoceptor-stimulated influx of 22Na+ into rat proximal tubules through the Na(+)-H+ exchanger was examined. The activity of this exchanger was defined as the component of 22Na+ uptake sensitive to inhibition by ethylisopropyl amiloride (EIPA) and was observed to be increased by both alpha 1- and alpha 2-adrenoceptor agonists as well as by phorbol 12-myristate 13-acetate (PMA). Selective alpha 2-adrenoceptor agonists produced a range of stimulation of EIPA-suppressible 22Na+ uptake: from a 72% increase above control with guanabenz to a 253% increase with B-HT 933. Because heterogeneity of alpha 2-adrenoceptor structure and function has been postulated, we examined whether the effects of alpha 2-adrenoceptors were sensitive to pertussis toxin. the responses to alpha 1-adrenoceptor agonists and PMA were unaffected, but the stimulation of Na(+)-H+ exchange by each of the selective alpha 2-adrenoceptor agonists tested was blocked. When Na(+)-H+ exchange was increased directly by PMA acting on protein kinase C, guanabenz but not B-HT 933 inhibited the response. The results indicated that the alpha 2-adrenoceptor agonists stimulated 22Na+ influx by activating a pertussis toxin-sensitive pathway but that certain alpha 2-adrenergic agonists such as guanabenz could additionally inhibit the exchanger through a pertussis toxin-resistant mechanism. This inhibition by guanabenz could be reversed by selective alpha 2-adrenoceptor antagonists.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Agonists↗

PubMed1990

Platelet-activating factor is a renal vasodilator in the anesthetized rat.

In view of the potent vasoactive properties of platelet-activating factor (PAF), we investigated the renal hemodynamic effects of this lipid. C16-PAF (0.5-10 ng/kg) given as a bolus into the renal arterial circulation of pentobarbital sodium-anesthetized male Wistar rats produced a dose-dependent increase in renal blood flow (6-15%), before causing systemic hypotension. The PAF-induced renal vasodilation and systemic hypotension was independent of renal innervation, unaltered by eicosanoid synthesis inhibition with indomethacin or dexamethasone, unchanged by the nonselective dopamine-receptor antagonist haloperidol, but was abolished by the PAF-receptor antagonist, L-659,989. Non-hypotensive intrarenal PAF infusion at 0.5 ng.min-1.kg-1 caused an increase in renal blood flow. Thus PAF can cause renal vasodilation in the rat kidney that is PAF-receptor mediated and does not involve the sympathetic nervous system or the release of vasodilatory arachidonic acid metabolites or dopamine.

Anesthesia↗

PubMed1989

Unmasking sensitive alpha 2-adrenoceptor-mediated renal vasoconstriction in conscious rats.

Renal vascular reactivity to intrarenal arterial boluses of norepinephrine (NE), phenylephrine (PHE; alpha 1-agonist), and guanabenz (GBZ; alpha 2-agonist) was assessed in conscious, freely moving, chronically instrumented, normotensive Wistar rats. Dose-response curves (DRCs) were obtained in the absence and cumulative presence of propranolol (PROP; beta-antagonist), corynanthine (CORY; alpha 1-antagonist) and idazoxan (IDX; alpha 2-antagonist) to estimate effective dosages (ED) required for 15 and 75% peak reductions in renal blood flow. The PHE DRC had a short shallow ED15 region, but was primarily linear with a steep slope. The GBZ DRC had a shallow slope. The NE DRC had a prolonged shallow phase in the ED15 region and a steep slope in the ED75 region. PROP had no effect on the DRCs. CORY caused a parallel rightward shift of the PHE DRC and had no effect on the GBZ DRC. After CORY, the shallow ED15 portion of the NE DRC was even more pronounced with a slope now identical to that of the GBZ DRC, whereas the ED75 region of the NE DRC was shifted rightward like the PHE DRC. IDX preferentially antagonized the ED15 regions of the GBZ and NE DRCs. In a second group of rats, the alpha 2-adrenoceptor antagonist, rauwolscine, was administered following base-line DRCs to demonstrate rightward shifts of the NE and GBZ DRCs when PHE DRCs remained unaffected. Therefore, when boluses of NE are injected into the rat kidney, the vasoconstrictive responses are a result of the activation of both alpha 1- and alpha 2-adrenoceptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Antagonists↗

PubMed1989

Differential interaction of guanabenz with receptor binding sites in rat brain and kidney.

The alpha 2-adrenoceptor selective agonist, [3H]guanabenz ([ 3H]GBZ), labels a unique population of binding sites in whole kidney which are not labeled by [3H]p-aminoclonidine ([3H]PAC). These binding sites are saturable and of high affinity (Kd = 10-12 nM). [3H]GBZ was not displaced from these sites by other alpha 1- or alpha 2-ligands, suggesting that they are non-adrenergic. This hypothesis is further supported by the insensitivity of renal guanabenz binding to regulation by guanyl nucleotides or to destruction by trypsin. Also, there appears to be no effect of guanabenz on the potency of isoproterenol in competing for beta-adrenoceptors in the kidney, which has been previously reported to be sensitive to clonidine. The absence of any effect of guanabenz on isoproterenol displacement of [3H]dihydroalprenolol in kidney suggests there are subtle differences in activation of alpha-receptors by clonidine and guanabenz in the kidney. In the brain, [3H]GBZ labels two binding sites. Part of the binding of [3H]GBZ in the brain is to sites essentially identical to the alpha 2-adrenoceptors labeled by [3H]PAC. The remainder of the binding resembles the non-adrenergic binding in kidney. The relationship of this unique binding site to the pharmacologic actions of guanabenz is currently not known.

Animals↗

PubMed1989

Alpha adrenoceptor agonist stimulation of oxygen consumption in rat proximal and distal nephrons.

Selective alpha-1 and alpha-2 adrenergic agonists were used to test the hypothesis that both receptor subtypes increase transcellular Na+ transport in the nephron. Oxygen consumption (QO2) was used as an index of transcellular transport and provided a continuous dynamic record of the tubules' response to an agonist. Both alpha-1 and alpha-2 adrenoceptor agonists produced a linear dose-related increase in QO2 at a steeper slope than the control in proximal and distal nephron segments. Stimulation of QO2 by the adrenergic agonists did not occur in the presence of ouabain and did not exceed the maximal respiratory rate achieved with nystatin. The response was determined to be a receptor-mediated increase in the ouabain-sensitive component of respiration. An inactive stereoisomer of epinephrine produced no effect, and adrenergic antagonists inhibited the stimulation by the respective agonists. Adrenergic agonists stimulated QO2 in proximal segments to a much greater degree than observed with suspensions of distal segments. These results are consistent with the density of adrenoceptors on nephrons reported in radioligand binding and autoradiographic studies. The alpha-1 agonists, cirazoline and phenylephrine, had similar dose-response curves and stimulated proximal tubules more than distal tubules. The alpha-2 agonists, guanabenz, UK 14,304 and B-HT 933, equivalently stimulated QO2 in distal tubules, but a spectrum of enhanced oxygen consumption was observed in proximal nephrons. Although both alpha adrenoceptor subtypes increased QO2 in proximal and distal nephrons, the mechanisms are likely to be different.

Adrenergic alpha-Agonists↗

PubMed1989

Synergistic alpha-1 and alpha-2 adrenergic stimulation of rat proximal nephron Na+/H+ exchange.

Both alpha-1 and alpha-2 adrenoceptors have been localized to the renal cortex, with the majority of binding sites on the proximal tubule. Because the major regulator of Na+ uptake into the proximal tubule is the Na+/H+ exchanger, and because alpha-1 and alpha-2 adrenoceptors stimulate it in other tissues, we tested the hypothesis that both alpha adrenoceptor subtypes can increase Na+ uptake into the proximal nephron by stimulating the Na+/H+ antiporter. Enhancement of Na+ transport by agonists was studied in isolated rat proximal tubules by determining the uptake of 22Na that was suppressible by the Na+/H+ inhibitor, 5-(N-ethyl-N-isopropyl)amiloride (EIPA). The phorbol ester, phorbol-12-myristate-13-acetate, (0.1 microM), directly stimulated the antiporter through protein kinase C and increased EIPA-suppressible 22Na uptake 250% above control. The alpha-1 adrenoceptor agonists, cirazoline and phenylephrine, in addition to the mixed agonist, norepinephrine, maximally stimulated uptake by 226 to 232% at 1 microM concentrations. alpha-2 agonists produced a range of maximal stimulations at 1 microM from 65% with guanabenz to 251% with B-HT 933. Increases in 22Na uptake by agonists were inhibited by selective adrenergic antagonists and by EIPA. The drugs did not change the EIPA-resistant component of 22Na uptake. Inasmuch as the adrenoceptor subtypes likely stimulated Na+/H+ exchange by differing intracellular pathways impinging upon common transport steps, we examined whether simultaneous stimulation of both pathways was additive. Submaximal concentrations (5 nM each) of alpha-1 and alpha-2 adrenoceptor agonists in combination synergistically enhanced 22Na uptake to a level similar to 1 microM concentrations of adrenoceptor agonists alone or in combination.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Agonists↗

PubMed1987

Improved separation method for rat proximal and distal renal tubules.

We modified and improved enzyme digestion and density gradient separation procedures to obtain fractions of proximal and distal renal tubules with high yield and viability. Kidneys from two anesthetized adult Wistar rats were flushed with Krebs-Henseleit buffer (KHB) and then perfused in situ with recirculated KHB containing collagenase and hyaluronidase at 125 mmHg. Cortices were excised, minced, and incubated in KHB containing enzymes for 35 min at 37 degrees C. Dissociated tubules were removed at 10-min intervals, rinsed, and placed in KHB containing 10% calf serum, vitamins, and amino acids at 4 degrees C. Separation was achieved by suspending the tissue in 45% isosmotic Percoll layered over an undiluted Percoll cushion and centrifuging. Proximal tubules sedimented near the cushion. Distal segments were isolated in the uppermost bands of a second 35% Percoll separation. Viability was greater than 95% as measured by lactate dehydrogenase leakage and quantitated by oxygen consumption and ATP content. Basal oxygen consumption was greater than 33 nmol O2 X min-1 X mg protein-1 in all fractions and was stimulated by succinate and inhibited by amiloride and ouabain. Basal ATP content averaged 9.7 nmol/mg ATP. An average 3.3-fold separation for the proximal fraction and 24.5-fold separation for the distal fraction was assessed by the enrichment of six specific enzyme markers, with several of the markers indicating separations up to 32-fold. Isolated tubules also displayed functional responses to parathyroid hormone and vasopressin. Distal, but not proximal, segments demonstrated significantly increased adenosine 3',5'-cyclic monophosphate formation with vasopressin.

Animals↗

PubMed1987

In vivo assessment of rat renal alpha adrenoceptors.

Despite a preponderance of alpha-2 adrenoceptors in homogenates of rat renal cortex, alpha-2 adrenoceptor agonists do not vasoconstrict the isolated buffer-perfused rat kidney. Both alpha-1 and alpha-2 adrenoceptor agonists can constrict the kidneys of dogs, cats and rabbits in vivo. Because alpha-2 adrenoceptor-mediated vasoconstriction is often difficult to demonstrate in vitro, and both subtypes of alpha agonists cause large increases in peripheral resistance in pithed rats, we tested the hypothesis that both alpha-1 and alpha-2 agonists would also constrict the rat kidney in vivo. Cannulation of the suprarenal artery and utilization of a high pressure liquid chromatography valve enabled random and reproducible intrarenal arterial bolus injections of agonists, and renal blood flow was monitored using Doppler flowmetry. Cirazoline, phenylephrine and norepinephrine bitartrate caused large renal vasopressor responses with minimal systemic effects. Although administered in a dosage range 100 to 1000 times that of alpha-1 agonists, the alpha-2 agonists (B-HT 920, UK 14,304 and guanabenz) produced only minimal renal vasoconstriction before systemic pressor effects. The low potency and efficacy of alpha-2 agonists could not be attributed to concomitant vasodilatory effects of these agents. Rat renal resistance vessels were less responsive to alpha-2 agonists than other species that have been examined. These studies are consistent with conclusions from in vitro examinations that only alpha-1 adrenoceptors mediate changes in renal vascular resistance and also autoradiographic studies reporting the localization of alpha-2 binding sites to rat renal tubules.

Adrenergic alpha-Agonists↗