Janusz Sadowski

An antihypertensive opioid: Biphalin, a synthetic non-addictive enkephalin analog decreases blood pressure in spontaneously hypertensive rats

BACKGROUND Endogenous opioid systems may be engaged in the control of arterial pressure (MAP), however, given the risk of addiction, opioid receptor agonists are not used in antihypertensive therapy. We examined cardiovascular effects of biphalin, a potentially non-addictive dimeric enkephalin analog, an agonist of opioid μ and δ receptors. METHODS Biphalin was infused iv at 150μg/kg/h to anesthetized spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). Along with MAP and heart rate (HR), renal blood flow (RBF) and iliac blood flow (IBF, a measure of hind limb perfusion) were measured using Transonic probes on renal and iliac artery, respectively. The effects of biphalin were compared with those of intravenous morphine (1.5mg/kg/h). RESULTS In two SHR groups biphalin decreased MAP from 143±2 to 130±2 and from 177±4 to 167±3mmHg (p<0.001) while HR did not change or modestly decreased. The renal blood flow (RBF) increased modestly and both renal and hind limb vascular resistances decreased significantly (p<0.001). The responses were blocked by inhibition of peripheral opioid receptors with naloxone methiodide. Unlike in SHR, in WKY rats biphalin did not change MAP or vascular resistances. Morphine infusion decreased MAP in SHR from 169±6 to 150±6mmHg (less decrease in WKY) and significantly decreased RBF and IBF. CONCLUSION Since biphalin, a non-addictive synthetic opioid, lowers MAP in SHR, a model of hypertension with pronounced neurogenic component, such analogs might find therapeutic application in human stress-induced hypertensive states. Biphalins advantage is no associated reduction of renal perfusion.

Intrarenal alterations of the angiotensin‐converting enzyme type 2/angiotensin 1‐7 complex of the renin‐angiotensin system do not alter the course of malignant hypertension in Cyp1a1‐Ren‐2 transgenic rats

The role of the intrarenal renin‐angiotensin system (RAS) in the pathophysiology of malignant hypertension is not fully understood. Accumulating evidence indicates that the recently discovered vasodilator axis of the RAS, angiotensin‐converting enzyme (ACE) type 2 (ACE2)/angiotensin 1–7 (ANG 1–7), constitutes an endogenous system counterbalancing the hypertensiogenic axis, ACE/angiotensin II (ANG II)/AT1 receptor. This study aimed to evaluate the role of the intrarenal vasodilator RAS axis in the pathophysiology of ANG II‐dependent malignant hypertension in Cyp1a1‐Ren‐2 transgenic rats. ANG II‐dependent malignant hypertension was induced by 13 days′ dietary administration of indole‐3‐carbinol (I3C), a natural xenobiotic that activates the mouse renin gene in Cyp1a1‐Ren‐2 transgenic rats. It was hypothesized that pharmacologically‐induced inhibition of the ACE2/ANG 1–7 complex should aggravate, and activation of this axis should attenuate, the course of ANG II‐dependent malignant hypertension. Blood pressure (BP) was monitored by radiotelemetry. ACE2 inhibitor (DX 600, 0.2 μg/day) and ACE2 activator (DIZE, 1 mg/day) were administrated via osmotic minipumps. Even though ACE2 inhibitor significantly decreased and ACE2 activator increased intrarenal ANG 1–7 concentrations, the course of BP, as well as of albuminuria, cardiac hypertrophy and renal glomerular damage, were not altered. It was shown that intrarenal alterations in the ACE2/ANG 1–7 complex did not significantly modify the course of malignant hypertension in I3C‐induced Cyp1a1‐Ren‐2 transgenic rats. Thus, in our experimental setting alterations of this intrarenal vasodilator complex of the RAS do not significantly modify the form of malignant hypertension that clearly depends on the inappropriately increased activity of the ACE/ANG II/AT1 receptor axis.

High salt intake increases blood pressure in normal rats: putative role of 20-HETE and no evidence on changes in renal vascular reactivity.

Background/Aims. High salt (HS) intake may elevate blood pressure (BP), also in animals without genetic salt sensitivity. The development of salt-dependent hypertension could be mediated by endogenous vasoactive agents; here we examined the role of vasodilator epoxyeicosatrienoic acids (EETs) and vasoconstrictor 20-hydroxyeicosatetraenoic acid (20-HETE). Methods. In conscious Wistar rats on HS diet systolic BP (SBP) was examined after chronic elevation of EETs using 4-[4-(3-adamantan-1-yl-ureido)-cyclohexyloxy]-benzoic acid (c-AUCB), a blocker of soluble epoxide hydrolase, or after inhibition of 20-HETE with 1-aminobenzotriazole (ABT). Thereafter, in acute experiments the responses of renal artery blood flow (Transonic probe) and renal regional perfusion (laser-Doppler) to intrarenal acetylcholine (ACh) or norepinephrine were determined. Results. HS diet increased urinary 20-HETE excretion. The SBP increase was not reduced by c-AUCB but prevented by ABT until day 5 of HS exposure. Renal vasomotor responses to ACh or norepinephrine were similar on standard and HS diet. ABT but not c-AUCB abolished the responses to ACh. Conclusions. 20-HETE seems to mediate the early-phase HS diet-induced BP increase while EETs are not engaged in the process. Since HS exposure did not alter renal vasodilator responses to Ach, endothelial dysfunction is not a critical factor in the mechanism of salt-induced blood pressure elevation.

Epoxyeicosatrienoic acid analog attenuates the development of malignant hypertension, but does not reverse it once established: A study in Cyp1a1-Ren-2 transgenic rats

Objective: We evaluated the therapeutic effectiveness of a new, orally active epoxyeicosatrienoic acid analog (EET-A) in rats with angiotensin II (ANG II)-dependent malignant hypertension. Methods: Malignant hypertension was induced in Cyp1a1-Ren-2 transgenic rats by activation of the renin gene using indole-3-carbinol (I3C), a natural xenobiotic. EET-A treatment was started either simultaneously with I3C induction process (early treatment) or 10 days later during established hypertension (late treatment). Blood pressure (BP) (radiotelemetry), indices of renal and cardiac injury, and plasma and kidney levels of the components of the renin–angiotensin system (RAS) were determined. Results: In I3C-induced hypertensive rats, early EET-A treatment attenuated BP increase (to 175 ± 3 versus 193 ± 4 mmHg, P < 0.05, on day 13), reduced albuminuria (15 ± 1 versus 28 ± 2 mg/24 h, P < 0.05), and cardiac hypertrophy as compared with untreated I3C-induced rats. This was associated with suppression of plasma and kidney ANG II levels (48 ± 6 versus 106 ± 9 and 122 ± 19 versus 346 ± 11 fmol ml−1 or g, respectively, P < 0.05) and increases in plasma and kidney angiotensin (1–7) concentrations (84 ± 9 versus 37 ± 6 and 199 ± 12 versus 68 ± 9 fmol/ml or g, respectively, P < 0.05). Remarkably, late EET-A treatment did not lower BP or improve renal and cardiac injury; indices of RAS activity were not affected. Conclusion: The new, orally active EET-A attenuated the development of experimental ANG II-dependent malignant hypertension, likely via suppression of the hypertensiogenic axis and augmentation of the vasodilatory/natriuretic axis of RAS.

Adenosine Effects on Renal Function in the Rat: Role of Sodium Intake and Cytochrome P450

Background/Aims: Adenosine (ADO) causes vasodilation in most tissues. In the kidney it can induce vasoconstriction or vasodilation, depending on the prevailing stimulation of A1 or A2 receptors (A1R, A2R). ADO-induced alterations of renal excretion may be secondary to haemodynamic changes, or reflect a direct influence on tubular transport. This whole-kidney study explored renal excretory responses to ADO receptor stimulation as related to renal haemodynamics sodium intake and cytochrome P450 (CYP-450) activity. Methods: The effects of ADO or an A2aR agonist (DPMA) on urine flow (V), sodium excretion (UNaV) and total solute excretion were examined in anaesthetized Wistar rats on a low-sodium or high-sodium (HS) diet. Total renal blood flow (RBF; renal artery probe), and outer- and inner-medullary blood flows (OM-BF, IM-BF; laser-Doppler fluxes) were also determined. Results: Consistent opposed effects of ADO and DPMA were only observed with the HS diet. ADO increased V (150%) and UNaV (100%); there were also significant increases in RBF, OM-BF and IM-BF. These changes were prevented by 1-aminobenzotriazol, a CYP-450 inhibitor. In HS rats, DPMA significantly decreased arterial blood pressure and renal excretion. Conclusions: Post-ADO diuresis/natriuresis was in part secondary to renal hyperperfusion; the response was probably mediated by CYP-450-dependent active agents. Selective A2aR stimulation induced systemic vasodilation, major hypotension, and a secondary decrease in renal excretion.

Progression of hypertension and kidney disease in aging fawn-hooded rats is mediated by enhanced influence of renin–angiotensin system and suppression of nitric oxide system and epoxyeicosanoids

ABSTRACT The fawn-hooded hypertensive (FHH) rat serves as a genetic model of spontaneous hypertension associated with glomerular hyperfiltration and proteinuria. However, the knowledge of the natural course of hypertension and kidney disease in FHH rats remains fragmentary and the underlying pathophysiological mechanisms are unclear. In this study, over the animals’ lifetime, we followed the survival rate, blood pressure (telemetry), indices of kidney damage, the activity of renin–angiotensin (RAS) and nitric oxide (NO) systems, and CYP450-epoxygenase products (EETs). Compared to normotensive controls, no elevation of plasma and renal RAS was observed in prehypertensive and hypertensive FHH rats; however, RAS inhibition significantly reduced systolic blood pressure (137 ± 9 to 116 ± 8, and 159 ± 8 to 126 ± 4 mmHg, respectively) and proteinuria (62 ± 2 to 37 ± 3, and 132 ± 8 to 87 ± 5 mg/day, respectively). Moreover, pharmacological RAS inhibition reduced angiotensin (ANG) II and increased ANG 1–7 in the kidney and thereby may have delayed the progression of kidney disease. Furthermore, renal NO and EETs declined in the aging FHH rats but not in the control strain. The present results, especially the demonstration of exaggerated vascular responsiveness to ANG II, indicate that RAS may contribute to the development of hypertension and kidney disease in FHH rats. The activity of factors opposing the development of hypertension and protecting the kidney declined with age in this model. Therefore, therapeutic enhancement of this activity besides RAS inhibition could be attempted in the therapy of human hypertension associated with kidney disease.

Vascular effects of a tripeptide fragment of novokinine in hypertensive rats: Mechanism of the hypotensive action

BACKGROUND Activation of angiotensin AT2 receptors (AT2R) counteracts vasoconstrictor effects of AT1R stimulation and contributes to blood pressure control. We examined effects on mean arterial pressure (MAP) and renal hemodynamics of LKP, a tripeptide fragment of novokinine, an established AT2R agonist. METHODS Effects of intravenous LKP infusion and then superimposed losartan (AT1R antagonist) on MAP, total renal (RBF, Transonic probe) and renal medullary blood flows (laser-Doppler), and on renal excretion, were examined in anesthetized (1) Wistar rats with acute norepinephrine-induced hypertension, untreated or pretreated with AT2R antagonist (PD 123319) and (2) spontaneously hypertensive rats (SHR) maintained on standard or high-sodium (HS) diet. RESULTS In Wistar rats LKP decreased MAP (-4%, p<0.01) and increased renal medullary perfusion, these effects were abolished in rats pretreated with PD 123319 in which a post-LKP increase in MAP (+6%, p<0.02) occurred. LKP did not alter MAP in SHR; in those on HS diet RBF decreased (-14%, p<0.02), the response that was reverted by losartan. Addition of losartan always decreased or tended to decrease MAP. CONCLUSIONS LKP lowered MAP in norepinephrine-induced hypertension, probably via activation of AT2R. At reduced availability of AT2R, as in SHR, LKP appeared to bind to different receptors, possibly AT1, and induced systemic or renal vasoconstriction. Compared to the parent novokinine, a smaller LKP molecule might be easier absorbed after oral application and more useful in therapy.

Interlobular Arteries From 2-Kidney, 1-Clip Goldblatt Hypertensive Rats' Exhibit-Impaired Vasodilator Response to Epoxyeicosatrienoic Acids

Background: Small renal arteries have a significant role in the regulation of renal hemodynamics and blood pressure (BP). To study potential changes in the regulation of vascular function in hypertension, we examined renal vasodilatory responses of small arteries from nonclipped kidneys of the 2‐kidney, 1‐clip Goldblatt hypertensive rats to native epoxyeicosatrienoic acids (EETs) that are believed to be involved in the regulation of renal vascular function and BP. A total of 2 newly synthesized EET analogues were also examined. Materials and Methods: Renal interlobular arteries isolated from the nonclipped kidneys on day 28 after clipping were preconstricted with phenylephrine, pressurized and the effects of a 14,15‐EET analogue, native 14,15‐EET and 11,12‐ether‐EET‐8ZE, an analogue of 11,12‐EET, on the vascular diameter were determined and compared to the responses of arteries from the kidneys of sham‐operated rats. Results: In the arteries from nonclipped kidneys isolated in the maintenance phase of Goldblatt hypertension, the maximal vasodilatory response to 14,15‐EET analogue was 30.1 ± 2.8% versus 49.8 ± 7.2% in sham‐operated rats; the respective values for 11,12‐ther‐EET‐8ZE were 31.4 ± 6.4% versus 80.4 ± 6%, and for native EETs they were 41.7 ± 6.6% versus 62.8 ± 4.4% (P ≤ 0.05 for each difference). Conclusions: We propose that reduced vasodilatory action and decreased intrarenal bioavailability of EETs combined with intrarenal angiotensin II levels that are inappropriately high for hypertensive rats underlie functional derangements of the nonclipped kidneys of 2‐kidney, 1‐clip Goldblatt hypertensive rats. These derangements could play an important role in pathophysiology of sustained BP elevation observed in this animal model of human renovascular hypertension.

Different blood pressure responses to opioids in 3 rat hypertension models: role of the baseline status of sympathetic and renin-angiotensin systems.

Opioids interact with sympathetic and renin-angiotensin systems in control of mean arterial pressure (MAP). Our earlier finding that biphalin, a synthetic enkephalin analogue, decreased MAP in anaesthetized spontaneously hypertensive rats (SHR) prompted us to further explore this action, to get new insights into pathogenesis of various forms of hypertension. Biphalin effects were studied in SHR, uninephrectomized rats on a high-salt diet (HS/UNX), and rats with angiotensin-induced hypertension (Ang-iH). Besides MAP, renal and iliac blood flows (RBF, IBF) and vascular resistances were measured. In anaesthetized and conscious SHR, biphalin (300 μg·h-1·kg-1 i.v.) decreased MAP by ∼10 and ∼20 mm Hg, respectively (P < 0.001). In anaesthetized HS/UNX and normotensive rats, MAP increased by ∼6-7 mm Hg (P < 0.02); without anaesthesia, only transient decreases occurred. MAP never changed in Ang-iH rats. Morphine (1.5 mg·h-1·kg-1 i.v.) decreased MAP in HS/UNX but only transiently so without anaesthesia; such anaesthesia dependence of response was also seen in normotensive rats. Ang-iH rats never responded to morphine. Hypotensive effect in SHR only depends primarily on the reduction by biphalin of vascular responsiveness to increased sympathetic stimulation; such increase is well documented for SHR. No MAP response to biphalin or morphine in Ang-iH could depend on angiotensin-induced alterations of the vascular wall morphology and function.

Sex-linked differences in the course of chronic kidney disease and congestive heart failure: a study in 5/6 nephrectomized Ren-2 transgenic hypertensive rats with volume overload induced using aorto-caval fistula.

The role of hypertension and the renin‐angiotensin system (RAS) in sex‐related differences in the course of chronic kidney disease (CKD) and congestive heart failure (CHF) remain unclear, especially when the two diseases are combined. In male and female Ren‐2 transgenic rats (TGR), a model of hypertension with activation of endogenous RAS, CKD was induced by 5/6 renal mass reduction (5/6 NX) and CHF was elicited by volume overload achieved by creation of an aorto‐caval fistula (ACF). The primary aim of the study was to examine long‐term CKD‐ and CHF‐related mortality, especially in animals with CKD and CHF combined, with particular interest in the potential sex‐related differences. The follow‐up period was 23 weeks after the first intervention (5/6 NX). We found, first, that TGR did not exhibit sexual dimorphism in the course of 5/6 NX‐induced CKD. Second, in contrast, TGR exhibited important sex‐related differences in the course of ACF‐induced CHF‐related mortality: intact female TGR showed higher survival rate than male TGR. This situation is reversed in the course of combined 5/6 NX‐induced CKD and ACF‐induced CHF‐related mortality: intact female TGR exhibited poorer survival than male TGR. Third, the survival rate in animals with combined 5/6 NX‐induced CKD and ACF‐induced CHF was significantly worsened as compared with rat groups that were exposed to ‘single organ disease’. Collectively, our present results clearly show that CKD aggravates long‐term mortality of animals with CHF. In addition, TGR exhibit remarkable sexual dimorphism with respect to CKD‐ and CHF‐related mortality, especially in animals with combined CKD and CHF.