A two-state receptor model for the interaction between angiotensin II type 1 receptors and non-peptide antagonists.

The interaction between non-peptide antagonists and the human angiotensin II type 1 (AT1) receptor in CHO-K1 cells was investigated by incubating the cells with antagonist, followed by a brief exposure to angiotensin II and measurement of the resulting inositol phosphate accumulation. The experimental data, expressed either as angiotensin II concentration-response curves or as antagonist concentration-inhibition curves, were in good agreement with computer-generated data according to a single-state model for the surmountable antagonist losartan and according to a two-step, two-state receptor model for the insurmountable antagonists candesartan, EXP3174, and irbesartan. Experimental and computer-generated data concerning the simultaneous exposure of the receptors to EXP3174 and losartan indicated that losartan produced a concentration-dependent restoration of the maximal response (angiotensin II concentration-response curves) as well as a rightward shift of the insurmountable portion of the EXP3174 inhibition curves, thus counteracting the higher-affinity EXP3174 binding. In conclusion, these findings provide further support for the concept that insurmountable and surmountable AT1 antagonists are mutually competitive and that insurmountable antagonist-receptor complexes may adopt different states.

How can the differences among AT1-receptor antagonists be explained?

Over the last few years we have seen a new class of antihypertensive drug evolve, the angiotensin II subtype 1 receptor antagonists. Hypothetically, all substances in this class should have the same effect on blood pressure and on end-organ damage as they all block the AT1 receptor. However, there are distinctions between them that may explain the significant and clinically important differences that seem to exist within this class of drug. An explanation for the differences may be found in receptor-antagonist kinetics. The receptor-antagonist interaction may be fitted to a two-state, two-step model which determines how large a part of the binding that will be surmountable and how large a part that will be insurmountable. The proportion of surmountable/insurmountable binding fits nicely to the duration of binding of the antagonist to the receptor, which may be translated into efficacy for the antagonist as outlined in the following review.

Impaired effect by NO synthase inhibition on tubuloglomerular feedback in rats after chronic renal denervation.

Acute unilateral renal denervation (aDNX) is associated with reduced tubuloglomerular feedback (TGF) sensitivity. Six days after denervation (cDNX) TGF sensitivity is somewhat restored, but TGF reactivity increased. This study aimed to investigate if the increased TGF reactivity that was seen in cDNX kidneys was owing to reduced production of nitric oxide (NO). TGF characteristics were determined with micropuncture experiments in anaesthetized rats, using the stop-flow pressure (PSF) technique. Maximal drop in PSF (DeltaPSF) was used as an index of TGF reactivity and the loop of Henle perfusion rate that elicited half-maximal DeltaPSF, the turning point (TP) was used as a measure of TGF sensitivity. In cDNX kidneys, TP was higher than in control rats (25.4 +/- 1.5 nL min-1 vs. 19.1 +/- 1.1 nL min-1), but clearly lower than in aDNX rats (37. 3 +/- 3.1 nL min-1). TGF was more reactive in cDNX rats (DeltaPSF=14. 7 +/- 1.1 mmHg) than in aDNX (7.9 +/- 1.1 mmHg) and control rats (9. 6 +/- 0.9 mmHg). Intratubular inhibition of NO synthase N omega-nitro-L-arginine (L-NA) in sham-DNX animals, decreased TP to 13.9 +/- 2.2 nL min-1 and DeltaPSF was increased with 92%. In cDNX kidneys TP was not significantly reduced by L-NA, and TGF reactivity was only moderately increased by 31%. Intratubular infusion of L-arginine (L-Arg) reduced DeltaPSF from 10.2 +/- 0.7 to 6.5 +/- 0.6 mmHg in sham-DNX kidneys, but TP was unaffected. In cDNX kidneys, there was no effect on either DeltaPSF or TP by the addition of L-Arg. However, when NO was delivered via sodium nitroprusside in the tubular perfusate, a clear reduction of DeltaPSF was seen in both sham-DNX and cDNX kidneys (from 9.9 +/- 0.5 to 4.4 +/- 1.0 and from14.9 +/- 1.3 to 8.1 +/- 1.5 mmHg, respectively). This indicates that cDNX is a state of low renal NO production and that this low level of NO resets TGF to a higher sensitivity and more pronounced reactivity.

Candesartan Causes Long-lasting Antagonism of the Angiotensin II Receptor-mediated Contractile Effects in Isolated Vascular Preparations: a Comparison with Irbesartan, Losartan and its Active Metabolite (EXP-3174).

Candesartan is a new angiotensin II type 1 (AT 1 ) receptor blocker. It displays insurmountable antagonism of angiotensin II responses, binding tightly to and dissociating slowly from the AT 1 -receptor. The purpose of this study was to compare the duration of angiotensin II antagonism by the AT 1 -receptor blockers candesartan, irbesartan, losartan and its active metabolite EXP-3174 in an isolated tissue preparation. The contractile response to angiotensin II was studied in the isolated portal vein of the rat, during incubation with AT1-receptor blockers and after an extensive washout period. The portal vein preparation was pre-stretched to a passive force of 5 mN in an organ bath filled with oxygenated Krebs' buffer at 37°C. The contractile tension developed by the vascular smooth muscles was monitored using a force-displacement transducer. The contractile response to repeated administration of angiotensin II was recorded before, during and following exposure (for 30-180 min) to candesartan, 0.1-1 nmol/l, irbesartan, 1-50 nmol/l, losartan, 30-100 nmol/l, and EXP-3174, 1-10 nmol/l. Drug exposure was followed by washing for up to 2 h. Candesartan produced a long-lasting blockade of the vascular contractile response to angiotensin II, as shown by maintenance of inhibition during the washout period. This effect of candesartan was independent of drug concentration and exposure time prior to washing. Irbesartan, losartan and EXP-3174 also blocked the angiotensin II-mediated contraction. However, in contrast to candesartan, the responses to angiotensin II rapidly returned towards baseline values during the washout period. The relatively short-lasting blockade by irbesartan, losartan and EXP-3174 was also independent of drug concentration and exposure time prior to washout. It is concluded that the AT 1 -receptor blockers differ in their ability to inhibit angiotensin II-mediated vascular contraction, with candesartan producing longer-lasting blockade than irbesartan, losartan and EXP-3174. The mechanism of the persistent inhibitory effect of candesartan is at present unclear. Possible explanations include tight binding and slow dissociation from the AT 1 -receptor, tissue accumulation resulting in 'local( dissociation and reassociation to the AT 1 -receptor, and stimulation of internalization of the AT 1 -receptor.

Mechanistic differences of various AT1-receptor blockers in isolated vessels of different origin.

The functional inhibitory characteristics of the angiotensin II type 1 receptor blockers (ARB) candesartan; irbesartan; and losartan and its active metabolite EXP 3174 (EXP) were studied in rabbit aortic strips and rat portal vein preparations in vitro. Moreover, plasma-protein binding was determined, and the binding was high (>98. 5%) for all ARBs. These values were needed to relate the concentrations of the ARBs used in vitro to the nonprotein bound concentrations in clinical use. In both vascular preparations, candesartan caused a marked decrease in the maximal contractile response of the angiotensin II (Ang II) concentration-response curve. Losartan, EXP, and irbesartan caused a rightward parallel shift without any major effects on the maximal response to Ang II. The inhibitory effect of candesartan developed slowly (maximal effect after >30 minutes) and lasted >2 hours despite repeated washing of the vessels. The effect of losartan, irbesartan, and EXP had a faster onset, and most of the inhibitory effect disappeared after washing. The duration of the inhibitory effects of the ARBs were not related to lipophilicity of the compounds. Cooling of the rat portal vein preparations to 4 degrees C before administration of candesartan prevented the persistent inhibition of Ang II response seen at 37 degrees C. For the other ARBs studied, the magnitude of inhibition and the speed of recovery of the Ang II response were independent of the incubation temperature before washing. In addition, when candesartan was given to conscious rats, the inhibitory effect on Ang II-induced blood pressure responses persisted during the 24-hour period despite nondetectable plasma concentrations of candesartan at 24 hours. It is concluded that functional inhibitory characteristics of candesartan differ from those of the other ARBs tested. At clinically relevant concentrations, candesartan is an insurmountable and long-lasting antagonist of the vascular contractile responses to Ang II.

Normalizing the expression of nitric oxide synthase by low-dose AT1 receptor antagonism parallels improved vascular morphology in hypertensive rats.

In essential hypertension, stroke and kidney damage may result from an impaired interaction of vasoregulatory systems. Stroke-prone spontaneously hypertensive rats (SHRSP) were studied to analyze the effects of a low-dose treatment of the angiotensin II type 1 receptor (AT1) blocker candesartan cilexetil on the expression of nitric oxide synthases (NOS) and on vascular structure. Both treated and untreated SHRSP were kept on a stroke-promoting dietary regimen, and compared with Wistar Kyoto rats (WKY). Early mortality of untreated SHRSP was prevented by the treatment. In untreated SHRSP, cerebral intraparenchymal vessels of the parietal lobe showed lesions of the vascular wall and its periphery, such as proteinaceous deposits, perivascular dilated spaces, increase in phagocytic cells, and decreased actin immunostaining. Renal lesions were more pronounced comprising arteriolar occlusion, extensive loss of actin, increased alpha1(IV) collagen expression, and glomerular sclerotic as well as tubulointerstitial lesions. Beneficial effects of the AT1 blockade were more pronounced in brain than in kidney. Activity profile of NOS showed increased NADPH diaphorase staining in media and endothelium of SHRSP; endothelial NOS3 immunoreactivity was decreased, but instead, inducible NOS2 increased in untreated SHRSP. These changes were largely prevented in the treated group. NOS activity in macula densa cells was unchanged, whereas afferent arteriolar renin levels were increased in untreated SHRSP. Results demonstrate an effective reduction of hypertensive vascular changes with a nonpressor dose of candesartan. A "role switch" of vascular NOS in hypertension from physiologic NOS3 toward deleterious NOS2 is suggested, and its prevention by the AT1 blocker points to an angiotensin II-dependent, nitric oxide-mediated pathway that may impair endothelial function and aggravate defects of the blood-brain barrier and kidney structures.

Combined treatment with ibuprofen and the AT1 receptor antagonist candesartan in young spontaneously hypertensive rats.

The current study was conducted to determine the potential influence of ibuprofen on the renal and systemic response to AT1 receptor blockade in conscious rats developing spontaneous hypertension. Experiments used spontaneously hypertensive rats (SHR) during the early developmental phase of hypertension (6 to 7 wk old). Six groups of rats were given the following during a 2-wk treatment protocol: (1) candesartan cilexetil (AT1 receptor antagonist) at 1 mg/kg body wt per d; (2) candesartan cilexetil at 10 mg/kg per d; (3) ibuprofen at 30 mg/kg per d; (4) a combination of candesartan cilexetil at 1 mg/kg per d + ibuprofen; (5) candesartan cilexetil at 10 mg/kg per d + ibuprofen; and (6) untreated (controls). All compounds were added to the drinking water at concentrations adjusted to maintain the desired dosage. In the young untreated SHR, systolic arterial pressure significantly increased from 134+/-4 to 170+/-11 mmHg. Candesartan at 1 mg/kg per d prevented any increase in arterial pressure (131+/-5 mmHg at week 0 versus 131+/-4 mmHg at week 2). At a dose of 10 mg/kg per d, candesartan lowered arterial pressure from 131+/-2 to 91+/-4 mmHg. Ibuprofen treatment alone had no effect on the increase in arterial pressure observed in young SHR over the study period, and had no effect on the changes produced by candesartan at either dose. In the two groups of rats receiving candesartan at 10 mg/kg per d (with and without ibuprofen), a significant increase in urine volume and water intake was observed; urine volume rose from 9.5+/-1.0 to 22.9+/-1.1 ml/d in rats given only candesartan and from 11.5+/-0.7 to 22.0+/-0.6 ml/d in rats given candesartan + ibuprofen. Urine volume and water intake were unchanged in all other groups. These effects on water handling are consistent with previous findings that chronic angiotensin II inhibition inhibits water reabsorption in the kidney. These results demonstrate that nonsteroidal anti-inflammatory drug treatment has no effect on the antihypertensive efficacy and diuretic effects of AT1 receptor blockade in rats developing hypertension.

Candesartan normalizes exaggerated tubuloglomerular feedback activity in young spontaneously hypertensive rats.

This study examines the effect of systemic blockade of angiotensin II AT1 receptors by candesartan on the exaggerated tubuloglomerular feedback (TGF) activity in 7-wk-old, euvolemic spontaneously hypertensive rats (SHR) and in Wistar-Kyoto rats (WKY). TGF activity was assessed by stop-flow pressure (SFP) and early proximal flow rate (EPFR) measurements during perfusion of Henle's loop. During the control period, SHR exhibited a greater maximal SFP response (19 versus 0.11 mmHg), and a lower tubular flow rate elicited half-maximal response (turning point) (12.7 versus 14.1 nl/min). In addition, EPFR at a high perfusion rate (40 nl/min) was lower in SHR, indicating exaggerated TGF activity. Blockade of AT1 receptors was achieved by intravenous injection of 0.05 mg/kg candesartan, which did not affect mean arterial pressure. Renal blood flow and mean arterial pressure responses to injections of angiotensin II were blocked by >95%. Maximum SFP response in SHR decreased to 11 mmHg, and turning point increased to 16.5 nl/min. The slope of the TGF response curve at the half-maximal SFP response (reactivity) decreased from -5.5 to -2.0 mmHg/nl per min. In contrast, maximum SFP response and TGF reactivity were unchanged by AT1 receptor blockade in euvolemic WKY. A small effect was noted as an increase in turning point to 18.0 nl/min after candesartan treatment. Thus, the exaggerated TGF activity in young SHR is markedly attenuated by systemic administration of candesartan, whereas TGF was basically unchanged in euvolemic WKY. These results demonstrate that angiotensin II plays an important role in the enhanced TGF activity observed in young SHR. Significant TGF activity, essentially at normal levels for euvolemic animals, persists during AT1 receptor blockade in both groups of rats.

Candesartan: a new-generation angiotensin II AT1 receptor blocker: pharmacology, antihypertensive efficacy, renal function, and renoprotection.

Candesartan, which is the active compound formed during adsorption of candesartan cilexetil, is one of the new generation of angiotensin II AT1 receptor blockers. Candesartan is an insurmountable blocker with a slow dissociation from the AT1 receptor, and it has been shown to effectively reduce BP in humans and in a variety of genetic and experimental models of hypertension. Possible mechanisms for a better effect in BP reduction compared with losartan may be the insurmountable characteristics of binding or more pronounced renal effects, but these need further evaluation. Candesartan has favorable effects on renal function demonstrated in both humans and animals, and has also been shown to protect the kidney in several models of renal injury. The beneficial effects exerted by candesartan could even be demonstrated in experimental models of hypertension in which BP is affected little, if at all. The renoprotective effects have been observed in the regulation of gene expression, as well as in biochemical and histologic evaluations.

11Beta-hydroxysteroid dehydrogenase, mineralocorticoid receptor, and thiazide-sensitive Na-Cl cotransporter expression by distal tubules.

Mineralocorticoid hormones regulate salt transport along the distal nephron by binding to intracellular receptors and activating gene transcription. Previous experiments showed that systemic aldosterone infusions stimulate thiazide-sensitive Na and Cl transport by distal convoluted tubule (DCT) cells; this effect could have been direct or secondary to systemic hormonal effects. Aldosterone target tissues express both mineralocorticoid receptors and the metabolic enzyme 11beta-hydroxysteroid dehydrogenase type 2. Mineralocorticoid receptors have been localized to the DCT in some experiments, but not in others. Expression of 11beta-hydroxysteroid dehydrogenase type 2 by DCT cells has not been investigated. The present experiments were designed to test the hypothesis that rat DCT cells are targets of aldosterone action. Patterns of mineralocorticoid receptor, 11beta-hydroxysteroid dehydrogenase, thiazide-sensitive Na-Cl cotransporter, and Na/Ca exchanger expression along the distal tubule were examined. A polyclonal antibody was generated to localize the thiazide-sensitive Na-Cl cotransporter. Thiazide-sensitive Na-Cl cotransporter and 11beta-hydroxysteroid dehydrogenase expression were examined using both in situ hybridization and immunocytochemistry; Na/Ca exchanger and mineralocorticoid receptor expression were examined by immunocytochemistry. The results indicate that 11beta-hydroxysteroid dehydrogenase is expressed by DCT cells, as well as connecting tubule cells and principal cells of the collecting duct; expression levels are low near the junction with the thick ascending limb and rise near the transition to the connecting tubule. Mineralocorticoid receptors are expressed by DCT cells, as well as along the thick ascending limb, connecting tubule, and collecting duct. The results indicate that components of the mineralocorticoid receptor system are expressed by DCT cells, suggesting that these cells are targets of aldosterone action.

Exaggerated tubuloglomerular feedback activity in genetic hypertension is mediated by ANG II and AT1 receptors.

The purpose of the present study was to determine the role of endogenous angiotensin II in exaggerated tubuloglomerular feedback (TGF) in young euvolemic spontaneously hypertensive rats (SHR). TGF was characterized by measuring proximal tubular stop-flow pressure (Psf) responses to loop of Henle perfusion before and during losartan infusion in 7-wk-old SHR and Wistar-Kyoto rats (WKY). In the control period, TGF responses were exaggerated in SHR compared with WKY. This was evidenced by a larger flow-induced maximum decrease in Psf (19 vs. 13 mmHg), lower turning point (8 vs.12 nl/min), and higher reactivity (-6.4 vs. -3.0 mmHg.nl-1.min-1) in SHR. Losartan (DuP-753) was infused into the renal artery to antagonize angiotensin AT1 receptors in the experimental period. This was verified by losartan inhibiting > 90% of the decrease in whole kidney and superficial cortical blood flow produced by exogenous angiotensin II in both strains. Losartan infusion significantly attenuated TGF activity in SHR but not in WKY. In SHR losartan reduced the maximum Psf response (from 19 to 10 mmHg) and increased the turning point (from 8 to 11 nl/min). SHR values during losartan administration were similar to those obtained in WKY. WKY values were unaffected by losartan. The lack of change in maximum TGF responses after losartan treatment was not unique to WKY, inasmuch as similar results were obtained in euvolemic Munich-Wistar rats (-2.0 +/- 0.7 and -1.1 +/- 1.0 mmHg vs. -8.4 +/- 0.7 mmHg in SHR). Thus angiotensin II does not appear to play an essential role in basal TGF activity during euvolemia in normotensive animals when there is minimal stimulation of the renin-angiotensin system. In contrast, our observations indicate that the exaggerated TGF in young euvolemic SHR represents a functional resetting that is dependent on angiotensin II and losartan-sensitive AT1 receptors during the development of genetic hypertension.

A new method for exchange of perfusion fluid during in vivo micropuncture.

The first step in the tubuloglomerular feedback (TGF) mechanism is postulated to be the sensing of changes in tubular NaCl concentration by the macula densa (MD) cells. Despite this, few in vivo studies using different tubular NaCl concentrations administered to the MD site have been completed. Methodological problems associated with retrograde perfusion might possibly explain this. In the present study we present a modification of a method used in in vitro tubular perfusion experiments which makes it possible to determine the TGF response using retrograde perfusion with a single perfusion pipette. The system is based on the use of a fluid exchange pipette introduced through a chamber in the holder of the perfusion pipette all the way to the tip of this pipette. The perfusion flow was regulated by a pressure head, and by regulating the outflow from the chamber. The flow was determined for different tip diameters and for different perfusion pressures. Using a tip diameter of 4 microns, the flow through the tip was in the range 18-25 nL min-1 at a pressure of 60-70 mmHg. The fluid-exchange pipette was tested in vivo by measuring proximal tubular stop-flow pressure while perfusing the macula densa region with different NaCl concentration.

Acute renal denervation causes time-dependent resetting of the tubuloglomerular feedback mechanism.

Renal effects of acute renal denervation (DNX) were studied in anaesthetized rats. In a first series, whole kidney clearance measurements were made 120 and 240 min after unilateral DNX. At 240 min, urine production was 3.59 +/- 0.87 microL min-1 in control kidneys and 7.74 +/- 1.97 microL min-1 in denervated kidneys. The corresponding values for sodium excretion were 0.56 +/- 0.17 and 1.41 +/- 0.34 mumol min-1, potassium excretion 0.48 +/- 0.08 and 0.97 +/- 0.37 mumol min-1 and glomerular filtration rate (GFR) 0.83 +/- 0.08 and 1.05 +/- 0.16 mL min-1, respectively. In a second series, tubuloglomerular feedback (TGF) characteristics were determined with the stop-flow pressure (Psf) technique. With increasing time, the sensitivity of the TGF mechanism diminished in denervated rats, as indicated by an increased turning point (TP). TP was significantly increased 2 h after DNX from 19.1 +/- 1.13 in control to 25.9 +/- 1.10 nL min-1. TP was further increased 4 h after DNX to 37.3 +/- 3.12 nL min-1. However, the maximal TGF response to increased flow in the late proximal tubule was not altered. But, Psf was significantly higher in DNX rats than in the controls (47.4 +/- 1.01 vs. 43.0 +/- 1.53 mmHg) in spite of a lower blood pressure (107 +/- 2.9 vs. 119 +/- 2.2 mmHg). We conclude that intact renal nerves are essential for the setting of the TGF sensitivity and hence the regulation of GFR.

Tubuloglomerular feedback in rats with chronic partial bilateral ureteral obstruction.

UNLABELLED: Rats with chronic partial unilateral ureteral obstruction (UUO) have a paradoxical resetting of the tubuloglomerular feedback (TGF) mechanism. During extracellular fluid volume expansion (VE) in control animals, a decrease in TGF sensitivity is normally noted. In rats with partial UUO, however, TGF sensitivity in the obstructed kidney was increased, associated with a relative reduction in single nephron glomerular filtration rate. In the present study, we examined the tubular and interstitial pressures, whole kidney function and the TGF system in rats with chronic partial bilateral ureteral obstruction. The rats were divided into preferentially ipsilaterally and preferentially contralaterally obstructed, as judged by pelvic volume. Measurements were performed both during hydropenia and during VE. During hydropenia TGF characteristics were the same in the 2 groups. During VE, however, TGF sensitivity was unchanged in the most obstructed kidneys (ipsilateral), while if the obstruction was preferentially contralateral, TGF-sensitivity decreased. This opposite change in TGF sensitivity resulted in higher electrolyte and water excretion from the least obstructed kidney. The change in TGF sensitivity was not correlated to renal interstitial pressure. IN CONCLUSION: rats with moderate chronic partial bilateral ureteral obstruction have an almost preserved function with regard to renal hemodynamics and excretion during hydropenia. During volume expansion, however, different responses were found in the least and most obstructed kidneys, with a decreased sensitivity of TGF in the least obstructed kidney, while TGF sensitivity was unchanged in the most obstructed kidney.

Resetting of tubuloglomerular feedback by interrupting early distal flow.

Cell-cell contact between the macula densa and the glomerular arterioles is is thought to provide the information pathway for the tubuloglomerular feedback (TGF) mechanism. When concentrations of sodium and chloride in the macula densa segment are increased, a signal is transmitted through the extraglomerular mesangium to contract the afferent arteriole. In addition, some observers have described a second region of contact between a later part of the distal tubule and the afferent arteriole of the same nephron. In this region the connecting tubule (CNT), and sometimes nerves that make contact with the cells of this CNT, were found. This arrangement gives another potential tubular segment, besides the macula densa plaque, in which the composition of tubular fluid may regulate glomerular dynamics. The present study was designed to investigate whether interrupting flow in the distal tubule downstream from the macula densa would influence the TGF mechanism. TGF was examined in rats by orthograde microperfusion, before and after blockade of the distal nephron with castor oil. Two variables were measured: maximum decrease in stop-flow pressure (delta Psf), and perfusion rate which elicits half-maximal decrease in delta Psf (V1/2). The fluid arriving at the blocking point was collected into a micro-pipette. The results show a significant increase in V1/2 from 19 to 25 nl min-1 after 30 min of blockade. In conclusion the results support a role of the distal nephron in the control of the TGF mechanism.

Tubule-tubule and tubule-arteriole contacts in rat kidney distal nephrons. A morphologic study based on computer-assisted three-dimensional reconstructions.

BACKGROUND: Functional investigations of the tubulo-glomerular feedback mechanism have indicated the existence of a contact between the distal nephron and the macula densa region. The structural justification of such a contact is investigated. EXPERIMENTAL DESIGN: Tubule-tubule and tubule-arteriole contacts were investigated in distal nephrons from normal rat kidneys. Computer-assisted three-dimensional reconstructions of distal nephrons were made from serial sections of renal cortical tissue and selected sections were examined by electron microscopy. RESULTS: In 14 of 15 reconstructed nephrons, the distal convoluted tubule or the connecting tubule approached the macula densa region. A wall-to-wall contact between two tubules corresponding to a three-dimensional distance below 28 microns between the axes of the two tubules was found in only five of the reconstructed tubules. The distal nephron contacts to afferent and efferent arterioles of the same nephron were also examined. The efferent arteriole revealed no consistent contacts but the afferent arteriole contacted the distal convoluted tubule/connecting tubules consistently in all 10 of the superficial nephrons and in 3 of 5 midcortical nephrons. Electron microscopy confirmed a close contact between the distal tubule and the afferent arteriole in superficial nephrons and small nerves were often found at or near the site of contact, but the morphology at the site of contact was not unique. The arteriole contacts were made with late distal convoluted tubules, connecting tubules, or cortical collecting ducts. CONCLUSIONS: In conclusion, the present study shows that tubule-tubule contacts are inconsistent between the macula densa region and the distal nephron but that the tubule-afferent arteriole contact is consistent and close in superficial nephrons. This morphology is compatible with the existence of a feedback mechanism between the superficial distal nephron and the afferent arteriole, apart from the one located at the juxtaglomerular apparatus.

Renal interstitial pressure and tubuloglomerular feedback control in rats during infusion of atrial natriuretic peptide (ANP).

Atrial natriuretic peptide (ANP), injected at physiological concentrations, is known to induce both natriuresis and diuresis. It has been suggested by some investigators that these changes result from an increasing glomerular filtration rate (GFR), but others have been unable to demonstrate an increased GFR. The tubuloglomerular feedback (TGF) mechanism is an important regulator of GFR, and the sensitivity of TGF is decreased during ANP administration. Furthermore, resetting of TGF is, in most instances, related to changes in renal interstitial hydrostatic and oncotic pressures. It is also known that ANP may increase capillary permeability which may change renal interstitial pressure. The present study was performed to examine renal interstitial pressures and the TGF mechanism during ANP infusion. In accordance with previous studies, TGF sensitivity was found to be decreased. The tubular flow rate which elicited half the maximal drop in stop-flow pressure (Psf) was increased from 18.5 to 25.7 nl min-1. In contrast, ANP infusion resulted in a decreased interstitial hydrostatic pressure and an increased interstitial oncotic pressure. From previous experiments, such changes in interstitial pressures would be expected to increase TGF sensitivity. The changes in interstitial pressure cannot, therefore, directly explain the resetting of the feedback mechanism. In conclusion, the present paper shows a decreased renal net interstitial pressure after intravenous administration of ANP.

Renal response to volume depletion and expansion in Milan hypertensive rats.

In previous studies on Milan hypertensive (MHS) rats, we found an impaired tubuloglomerular feedback (TGF) response before, during and after development of hypertension. In the present study MHS rats and rats of the Milan normotensive strain (MNS) were investigated after 24 hours of volume depletion (VD) and subsequently after 5% isotonic volume expansion (VE) with respect to whole kidney function, interstitial hydrostatic (P(int)) and oncotic (IIint) pressures, stop-flow pressure characteristics of TGF and changes in early proximal flow rate in response to increased loop of Henle flow. MHS rats had higher mean arterial blood pressure (Pa) than MNS rats (129 vs. 101 mmHg) both after VD and after subsequent VE. No difference in glomerular filtration rate (GFR) was found. Both strains had a low urine flow rate (approximately 1.5 microliters min-1) during VD, which increased fourfold after VE. The interstitium was significantly more dehydrated in MHS, as indicated by a more negative net interstitial pressure (P(int)-IIint than in MNS (-1.3 +/- 0.3 vs. +/- 0.0 +/- 0.5 mmHg) after VE. The TGF mechanism was more activated in MHS during volume depletion, as indicated by a larger drop in stop-flow pressure (Psf) in response to loop of Henle perfusion (7.1 +/- 0.7 vs. 4.7 +/- 0.2 mmHg, P less than 0.05). However, during VD the loop of Henle flow that elicited half maximal response in Psf, the turning point (TP), was equally low in MHS and MNS (13.5 +/- 0.6 and 14.3 +/- 0.4, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)