Sex Dependence in Control of Renal Haemodynamics and Excretion in Streptozotocin Diabetic Rats-Role of Adenosine System and Nitric Oxide.

Recently, we compared an interplay of the adenosine system and nitric oxide (NO) in the regulation of renal function between male normoglycaemic (NG) and streptozotocin-induced diabetic rats (DM). Considering the between-sex functional differences, e.g., in the NO status, we present similar studies performed in female rats. We examined if the theophylline effects (non-selective adenosine antagonist) in NG and DM females with or without active NO synthases differed from the earlier findings. In anaesthetised female Sprague Dawley rats, both NG and DM, untreated or after NO synthesis blockade with L-NAME, theophylline effects, on blood pressure, renal hemodynamics and excretion, and renal tissue NO were investigated. Renal artery blood flow (Transonic probe), cortical, outer-, and inner-medullary flows (laser-Doppler technique), and renal tissue NO signal (selective electrode) were measured. In contrast to males, in female NG and DM rats, theophylline induced renal vasodilation. In NO-deficient females, theophylline induced comparable renal vasodilatation, confirming the vasoconstrictor influence of the renal adenosine. In NG and DM females with intact NO synthesis, adenosine inhibition diminished kidney tissue NO, contrasting with an increase reported in males. Lowered baseline renal excretion in DM females suggested stimulation of renal tubular reabsorption due to the prevalence of antinatriuretic over natriuretic tubular action of adenosine receptors. An opposite inter-receptor balance pattern emerged previously from male studies. The study exposed between-sex functional differences in the interrelation of adenosine and NO in rats with normoglycaemia and streptozotocin diabetes. The findings also suggest that in diabetes mellitus, the abundance of individual receptor types can distinctly differ between females and males.

Nonselective and A2a-Selective Inhibition of Adenosine Receptors Modulates Renal Perfusion and Excretion Depending on the Duration of Streptozotocin-Induced Diabetes in Rats.

Long-lasting hyperglycaemia may alter the role of adenosine-dependent receptors (P1R) in the control of kidney function. We investigated how P1R activity affects renal circulation and excretion in diabetic (DM) and normoglycaemic (NG) rats; the receptors' interactions with bioavailable NO and H2O2 were also explored. The effects of adenosine deaminase (ADA, nonselective P1R inhibitor) and P1A2a-R-selective antagonist (CSC) were examined in anaesthetised rats, both after short-lasting (2-weeks, DM-14) and established (8-weeks, DM-60) streptozotocin-induced hyperglycaemia, and in normoglycaemic age-matched animals (NG-14, NG-60, respectively). The arterial blood pressure, perfusion of the whole kidney and its regions (cortex, outer-, and inner medulla), and renal excretion were determined, along with the in situ renal tissue NO and H2O2 signals (selective electrodes). The ADA treatment helped to assess the P1R-dependent difference in intrarenal baseline vascular tone (vasodilation in DM and vasoconstriction in NG rats), with the difference being more pronounced between DM-60 and NG-60 animals. The CSC treatment showed that in DM-60 rats, A2aR-dependent vasodilator tone was modified differently in individual kidney zones. Renal excretion studies after the ADA and CSC treatments showed that the balance of the opposing effects of A2aRs and other P1Rs on tubular transport, seen in the initial phase, was lost in established hyperglycaemia. Regardless of the duration of the diabetes, we observed a tonic effect of A2aR activity on NO bioavailability. Dissimilarly, the involvement of P1R in tissue production of H2O2, observed in normoglycaemia, decreased. Our functional study provides new information on the changing interaction of adenosine in the kidney, as well as its receptors and NO and H2O2, in the course of streptozotocin diabetes.

Role of Ang1-7 in renal haemodynamics and excretion in streptozotocin diabetic rats.

The contribution of angiotensin (1-7) (Ang1-7) to control of extrarenal and renal function may be modified in diabetes. We investigated the effects of Ang1-7 supplementation on blood pressure, renal circulation and intrarenal reactivity (IVR) to vasoactive agents in normoglycaemic (NG) and streptozotocin diabetic rats (DM). In Sprague Dawley DM and NG rats, 3 weeks after streptozotocin (60 mg/kg i.p.) or solvent injection, Ang1-7 was administered (400 ng/min) over the next 2 weeks using subcutaneously implanted osmotic minipumps. For a period of 5 weeks, blood pressure (BP), 24 h water intake and diuresis were determined weekly. In anaesthetised rats, BP, renal total and cortical (CBF), outer (OMBF) and inner medullary (IMBF) perfusion and urine excretion were determined. To check IVR, a short-time infusion of acetylcholine or norepinephrine was randomly given to the renal artery. Unexpectedly, BP did not differ between NG and DM, and this was not modified by Ang-1-7 supplementation. Baseline IMBF was higher in NG vs. DM, and Ang1-7 treatment did not change it in NG but decreased it in DM. In the latter, Ang1-7 increased cortical IVR to vasoconstrictor and vasodilator stimuli. IMBF decrease after high acetylcholine dose seen in untreated NG was reverted to an increase in Ang1-7 treated rats. Irrespective of the glycaemia level, Ang1-7 did not modify BP. However, it impaired medullary circulation in DM, whereas in NG it rendered the medullary vasculature more sensitive to vasodilators. Possibly, the medullary hypoperfusion in DM was mediated by Ang1-7 activation of angiotensin AT-1 receptors which are upregulated by hyperglycaemia.

Interplay of the adenosine system and NO in control of renal haemodynamics and excretion: Comparison of normoglycaemic and streptozotocin diabetic rats.

The adenosine (Ado) system may participate in regulation of kidney function in diabetes mellitus (DM), therefore we explored its role and interrelation with NO in the control of renal circulation and excretion in normoglycemic (NG) and streptozotocin-diabetic (DM) rats. Effects of theophylline (Theo), a non-selective Ado receptor antagonist, were examined in anaesthetized NG or in streptozotocin induced diabetic (DM) rats, untreated or after blockade of NO synthesis with l-NAME. We measured arterial blood pressure (MABP), whole kidney blood flow and renal regional flows: cortical and outer- and inner-medullary (IMBF), determined as laser-Doppler fluxes. Renal excretion of water, total solutes and sodium and in situ renal tissue NO signal (selective electrodes) were also determined. Theo experiments disclosed minor baseline vasoconstrictor and vasodilator tone in the kidney of NG and DM rats, respectively. NO blockade increased baseline MABP and decreased renal haemodynamics, similarly in NG and DM rats, indicating comparable vasodilator influence of NO in the two groups. Unexpectedly, in all rats with intact NO synthesis, Ado receptor blockade increased kidney tissue NO. In NO-deficient NG and DM rats, Ado receptor blockade induced comparable renal vasodilatation, suggesting similar vasoconstrictor influence of the Ado system. However, DM rats showed an unexplained association of decreased MABP and IMBF and increased NO signal. Higher baseline renal excretion in DM rats indicated inhibition of renal tubular reabsorption due to the prevalence of natriuretic A2 over antinatriuretic A1 receptors. In conclusion, the experiments provided new insights in functional interrelation of adenosine and NO in normoglycaemia and streptozotocin-diabetes.

TMAO, a seafood-derived molecule, produces diuresis and reduces mortality in heart failure rats.

Trimethylamine-oxide (TMAO) is present in seafood which is considered to be beneficial for health. Deep-water animals accumulate TMAO to protect proteins, such as lactate dehydrogenase (LDH), against hydrostatic pressure stress (HPS). We hypothesized that TMAO exerts beneficial effects on the circulatory system and protects cardiac LDH exposed to HPS produced by the contracting heart. Male, Sprague-Dawley and Spontaneously-Hypertensive-Heart-Failure (SHHF) rats were treated orally with either water (control) or TMAO. In vitro, LDH with or without TMAO was exposed to HPS and was evaluated using fluorescence correlation spectroscopy. TMAO-treated rats showed higher diuresis and natriuresis, lower arterial pressure and plasma NT-proBNP. Survival in SHHF-control was 66% vs 100% in SHHF-TMAO. In vitro, exposure of LDH to HPS with or without TMAO did not affect protein structure. In conclusion, TMAO reduced mortality in SHHF, which was associated with diuretic, natriuretic and hypotensive effects. HPS and TMAO did not affect LDH protein structure.

Heart failure is a common cause of death in industrialized countries with aging populations. Japan, however, has lower rates of heart failure and fewer deaths linked to this disease than the United States or Europe, despite having the highest proportion of elderly people in the world. Dietary differences between these regions may explain the lower rate of heart failure in Japan. The Japanese diet is rich in seafood, which contains nutrients that promote heart health, such as omega-3 fatty acids. Seafood also contains other compounds, including trimethylamine oxide (TMAO). Fish that live in deep waters undergo high pressures, which can damage their proteins, but TMAO seems to protect the proteins from harm. In humans, eating seafood increases TMAO levels in the blood and urine, but it is unclear what effects this has on heart health. Increased levels of TMAO in the blood are associated with cardiovascular diseases, but scientists are not sure whether TMAO itself harms the heart. A toxic byproduct of gut bacteria called TMA is converted in TMAO in the body, so it is possible that TMA rather than TMAO is to blame. To assess the effects of dietary TMAO on heart failure, Gawrys-Kopczynska et al. fed the compound to healthy rats and rats with heart failure for one year. TMAO had no effects on the healthy rats. Of the rats with heart failure that were fed TMAO, all of them survived the year, while one third of rats with heart failure that were not fed TMAO died. TMAO-treated rats with heart failure had lower blood pressure and urinated more than untreated rats with the condition. The experiments suggest that dietary TMAO may mimic the effects of heart failure treatments, which remove excess water and salt and lower pressure on the heart. More studies are needed to confirm whether TMAO has this same effect on humans.

Clopidogrel Partially Counteracts Adenosine-5'-Diphosphate Effects on Blood Pressure and Renal Hemodynamics and Excretion in Rats.

BACKGROUND: Adenosine-5'-diphosphate (ADP) can influence intrarenal vascular tone and tubular transport, partly through activation of purine P2Y12 receptors (P2Y12-R), but their actual in vivo role in regulation of renal circulation and excretion remains unclear. METHODS: The effects of intravenous ADP infusions of 2-8mg/kg/hour were examined in anesthetized Wistar rats that were untreated or chronically pretreated with clopidogrel, 20mg/kg/24hours, a selective P2Y12-R antagonist. Renal blood flow (transonic probe) and perfusion of the superficial cortex and medulla (laser-Doppler fluxes) were measured, together with urine osmolality (Uosm), diuresis (V), total solute (UosmV), sodium (UNaV) and potassium (UKV) excretion. RESULTS: ADP induced a gradual, dose-dependent 15% decrease of mean arterial pressure, a sustained increase of renal blood flow and a 25% decrease in renal vascular resistance. Clopidogrel pretreatment attenuated the mean arterial pressure decrease, and did not significantly alter renal blood flow or renal vascular resistance. Renal medullary perfusion was not affected by ADP whereas Uosm decreased from 1,080 ± 125 to 685 ± 75 mosmol/kg H20. There were also substantial significant decreases in UosmV, UNaV and UKV; all these changes were attenuated or abolished by clopidogrel pretreatment. Two-weeks' clopidogrel treatment decreased V while UosmUosmV and UNaV increased, most distinctly after 7 days. Acute clopidogrel infusion modestly decreased mean arterial pressure and significantly increased outer- and decreased inner-medullary perfusion. CONCLUSIONS: Our functional studies show that ADP can cause systemic and renal vasodilation and a decrease in mean arterial pressure, an action at least partly mediated by P2Y12 receptors. We confirmed that these receptors exert tonic action to reduce tubular water reabsorption and urine concentration.

Hypertension in rats is associated with an increased permeability of the colon to TMA, a gut bacteria metabolite.

An increased blood trimethylamine N-oxide (TMAO) has emerged as a marker of cardiovascular mortality, however, the mechanisms of the increase are not clear. We evaluated if hypertension was associated with changes in the colon permeability to trimethylamine (TMA), a TMAO precursor. We did experiments on male, 24-26-week-old normotensive Wistar-Kyoto rats (WKY), spontaneously hypertensive rats (SHR) and SHR treated with enalapril, an antihypertensive drug (SHR-E). To check the colon permeability and liver TMA clearance, blood was collected from the portal vein and hepatic veins confluence, at baseline and after the intracolonic administration of TMA. Arterial blood pressure (BP) and intestinal blood flow (IBF) recordings and histological assessment of the colon were performed. SHR showed an increased gut-blood barrier permeability to TMA. Namely, at baseline SHR had a higher BP and portal blood TMA, but a lower IBF than WKY. After the intracolonic administration of TMA, SHR had a significantly higher portal blood TMA and higher TMA liver clearance than WKY. In SHR the arteriolar walls of the colon mucosa were significantly thicker than in WKY. Furthermore, SHR showed a significant decrease in the height of the mucosa. In contrast, SHR-E had lower portal blood TMA, lower BP and smaller thickness of arteriolar walls, but higher IBF than SHR, which indicates improved function of the gut-blood barrier in SHR-E. All groups had similar immunostaining of occludin and zonula occludens-1, markers of tight junctions. In conclusion, hypertensive rats show an increased permeability of the colon to TMA, which is accompanied by morphological and hemodynamic alterations in the colon. Therefore, cardiovascular diseases may be characterized by an increased permeability of the gut-blood barrier to bacterial metabolites such as TMA.

Intracolonic hydrogen sulfide lowers blood pressure in rats.

Research suggests that hydrogen sulfide (H2S) is an important biological mediator involved in various physiological processes including the regulation of arterial blood pressure (BP). Although H2S is abundant in the colon, the effects of gut-derived H2S on the circulatory system have not yet been investigated. We studied the effects of intracolonic administration of Na2S, a H2S donor, on systemic hemodynamics. Hemodynamics were recorded in anesthetized, normotensive Wistar Kyoto and spontaneously hypertensive rats at baseline and after intracolonic injection of either saline (controls) or Na2S·9H2O saline solution at a dose range of 10-300 mg/kg of BW. The H2S donor produced a significant, dose-dependent decrease in mean arterial blood pressure (MABP), which lasted several times longer than previously reported after parenteral infusions (>90 min). The effect was more pronounced in hypertensive than in normotensive rats. The Na2S-induced decrease in MABP was reduced by pretreatment with glibenclamide, an inhibitor of ATP-sensitive potassium-channels. Na2S did not affect mesenteric vein blood flow. Rats treated with Na2S showed increased portal blood levels of thiosulfate and sulfane sulfur, products of H2S oxidation. In contrast, rats treated with neomycin, an antibiotic, showed significantly decreased levels of thiosulfate and sulfane sulfur, and a tendency for greater hypotensive response to Na2S. The H2S donor decreased heart rate but did not affect ECG morphology and QTc interval. In conclusion the gut-derived H2S may contribute to the control of BP and may be one of the links between gut microbiota and hypertension. Furthermore, gut-derived H2S may be a therapeutic target in hypertension.

Influence of P2X receptors on renal medullary circulation is not altered by angiotensin II pretreatment.

BACKGROUND: Purine P2X and P2Y receptors (P2-R) are involved in control of renal circulation, especially in the medulla, wherein they appear to interact with angiotensin II (Ang II). Our experimental approach enabled examination of interaction with Ang II per se, in the absence of blood pressure elevation. In this whole-kidney functional study we focused on the influence of P2X1-R on perfusion of the renal medulla. METHODS: Acute experiments were conducted with normal rats, untreated or subjected to two weeks' infusion of Ang II (osmotic minipumps). Urethane was used for anesthesia because in Ang II-treated rats it normalized elevated blood pressure. MRS2159, a P2X1-R inhibitor, was infused intravenously or directly into the medulla. Renal blood flow (RBF, Transonic renal artery probe), perfusion of the outer and inner medulla (OM-BF, IM-BF; laser-Doppler fluxes), and sodium and water excretion and urine osmolality (Uosm) were measured. RESULTS: In untreated rats intravenous MRS2159 unexpectedly decreased RBF by 12±4% (p<0.02) and IM-BF by 7±2% (p<0.05). In Ang II-pretreated rats the inhibitor tended to increase RBF while OM-BF and IM-BF increased 14±5% and 12±2%, respectively (p<0.05 for both). Renal excretion was not affected, with or without Ang II treatment, while Uosm increased by about 150mosmol/kg H2O (p<0.05). Intramedullary MRS2159 increased IM-BF only, by 21±5% in untreated and 16±3% in Ang II-treated rats (p<0.04 for both). CONCLUSIONS: Tonic activity of P2X1 receptors participates in control of renal medullary perfusion and of the tubular processes involved in urine concentration, neither effect is modified by Ang II pretreatment.

Role of atrial natriuretic peptide in mediating the blood pressure-independent natriuresis elicited by systemic inhibition of nitric oxide.

While it is clearly recognized that increased intrarenal nitric oxide (NO) levels elicit natriuresis, confounding data showing that systemic nitric oxide synthase inhibition (NOSi) also increases sodium excretion (UNaV) poses a conundrum. This response has been attributed to the associated increases in arterial pressure (AP); however, the increases in AP and in UNaV are temporally dissociated. The changes in regional renal haemodynamics induced by NOSi could also contribute to the alterations of UNaV. To evaluate the roles of AP and non-AP mechanisms mediating the natriuresis, N ω-nitro-L-arginine methyl ester hydrochloride (L-NAME) was infused i.v. at doses ranging from 5 to 50 µg/kg/min in anaesthetized rats. UNaV, perfusion of the cortex (cortical blood flow, CBF) and medulla (medullary blood flow, MBF) with laser-Doppler flowmetry and glomerular filtration rate (GFR) were measured. UNaV increased from 0.6 ± 0.2 to 1.6 ± 0.1 µmol/kg/min (P < 0.05) with the lower nonpressor doses. With the higher doses, AP increased from 116 ± 4 to 122 ± 4 mmHg and UNaV increased from 1.1 ± 0.3 to 3.3 ± 0.7 µmol/min/g (P < 0.002). UNaV increased similarly in a group where renal AP was maintained at baseline levels. The associated reductions in CBF (17 ± 5 and 38 ± 5 %) and MBF (27 ± 6 and 52 ± 6 %) would be expected to attenuate rather than contribute to the natriuresis. Plasma atrial natriuretic peptide (ANP) concentrations increased significantly following NOSi. Anantin, a natriuretic peptide receptor-A blocker, prevented or reversed the L-NAME-induced natriuresis without altering the L-NAME-induced changes in AP or CBF. The results indicate that increased ANP and related natriuretic peptides mediate the AP-independent natriuresis, at least partly, elicited by systemic L-NAME infusion and help resolve the conundrum of natriuresis during systemic NOSi.

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.

Renal heme oxygenase-1 induction with hemin augments renal hemodynamics, renal autoregulation, and excretory function.

Heme oxygenases (HO-1; HO-2) catalyze conversion of heme to free iron, carbon monoxide, and biliverdin/bilirubin. To determine the effects of renal HO-1 induction on blood pressure and renal function, normal control rats (n = 7) and hemin-treated rats (n = 6) were studied. Renal clearance studies were performed on anesthetized rats to assess renal function; renal blood flow (RBF) was measured using a transonic flow probe placed around the left renal artery. Hemin treatment significantly induced renal HO-1. Mean arterial pressure and heart rate were not different (115 ± 5 mmHg versus 112 ± 4 mmHg and 331 ± 16 versus 346 ± 10 bpm). However, RBF was significantly higher (9.1 ± 0.8 versus 7.0 ± 0.5 mL/min/g, P < 0.05), and renal vascular resistance was significantly lower (13.0 ± 0.9 versus 16.6 ± 1.4 [mmHg/(mL/min/g)], P < 0.05). Likewise, glomerular filtration rate was significantly elevated (1.4 ± 0.2 versus 1.0 ± 0.1 mL/min/g, P < 0.05), and urine flow and sodium excretion were also higher (18.9 ± 3.9 versus 8.2 ± 1.0 µL/min/g, P < 0.05 and 1.9 ± 0.6 versus 0.2 ± 0.1 µmol/min/g, P < 0.05, resp.). The plateau of the autoregulation relationship was elevated, and renal vascular responses to acute angiotensin II infusion were attenuated in hemin-treated rats reflecting the vasodilatory effect of HO-1 induction. We conclude that renal HO-1 induction augments renal function which may contribute to the antihypertensive effects of HO-1 induction observed in hypertension models.

The renin-angiotensin system and the third mechanism of renal blood flow autoregulation.

Autoregulation of renal blood flow comprises three mechanisms: the myogenic response (MR), the tubuloglomerular feedback (TGF), and a third mechanism (3M). The nature of 3M is unknown; it may be related to hypotensive resetting of autoregulation that probably relies on pressure-dependent stimulation of the renin-angiotensin system (RAS). Thus we used a normotensive angiotensin II clamp in anesthetized rats and studied autoregulation 1) by slow ramp-shaped reductions in renal perfusion pressure (RPP) followed by ramp-shaped RPP restorations and 2) by means of the step response technique: after 30 s of either total or partial suprarenal aortic occlusion, a step increase in RPP was made and the response of renal vascular conductance analyzed to assess the mechanisms' strength and initial direction (vasodilation or constriction). The angiotensin clamp abolished the resetting of autoregulation during ramp-shaped RPP changes. Under control conditions, the initial TGF response was dilatory after total occlusions but constrictive after partial occlusions. The initial 3M response presented a mirror image to the TGF: it was constrictive after total but dilatory after partial occlusions. The angiotensin clamp suppressed the TGF and turned the initial 3M response following total occlusions into dilation. We conclude that 1) pressure-dependent RAS stimulation is a major cause behind hypotensive resetting of autoregulation, 2) TGF sensitivity strongly depends on pressure-dependent changes in RAS activity, 3) the 3M is modulated, but not mediated, by the RAS, and 4) the 3M acts as a counterbalance to the TGF and might possibly be related to the recently described connecting tubule glomerular feedback.

Sodium intake determines the role of adenosine A2 receptors in control of renal medullary perfusion in the rat.

BACKGROUND: In the kidney, adenosine (ADO) can induce either vasoconstriction or vasodilatation, mediated by A1 or A2 receptors, respectively. The vasodilator influence may be of special importance in the renal medulla which operates at low tissue pO(2) levels and is susceptible to ischaemic damage. It has not been established if ADO induced vasodilatation is modified by salt intake. METHODS: We examined effects of stimulation or inhibition of ADO receptors (A2R) on perfusion of the renal cortex and medulla on low- or high- sodium intake (LS, HS). Effects of suprarenal aortic ADO (0.03 mmol/kg/h), A2R agonist (DPMA), 0.08-0.4 mmol/kg/h, or antagonist (DMPX), 1.7 micromol/kg/h, were examined in anaesthetized rats maintained on LS (0.15% Na) or HS (4% Na) diet for 3 weeks. Whole kidney blood flow (RBF) and the perfusion (laser-Doppler) of the superficial cortex and outer and inner medulla (OM-BF, IM-BF) were measured. RESULTS: In LS rats neither drug changed renal perfusion. In HS rats ADO increased RBF 18 +/- 3%, OM-BF 16 +/- 7% and IM-BF 16 +/- 6%. IM-BF increased after DPMA 18 +/- 5% and decreased after DMPX 13 +/- 3%; neither drug consistently changed perfusion of the cortex. CONCLUSIONS: On HS intake, medullary perfusion is controlled by ADO vasodilator (A2) receptors, which may help provide adequate oxygen to the medulla, the zone which normally operates under relative hypoxia. On LS intake, the vasodilator and vasoconstrictor effects are probably in balance and ADO has little role in control of intrarenal circulation.

Role of NO and COX pathways in mediation of adenosine A1 receptor-induced renal vasoconstriction.

The mechanism of adenosine A1 receptor-induced intrarenal vasoconstriction is unclear; it depends on sodium intake and may be mediated by changing the intrarenal activity of the nitric oxide (NO) and/or cyclooxygenase (COX) pathway of arachidonic acid metabolism. The effects of 2-chloro-N(6)-cyclopentyl-adenosine (CCPA), a selective A1 receptor agonist, on renal hemodynamics were examined in anesthetized rats maintained on high sodium (HS) or low sodium (LS) diet. Total renal (i.e., cortical) blood flow (RBF) as well as superficial cortical (CBF), outer medullary (OMBF), and inner medullary (IMBF) flows were determined by laser-Doppler. In HS rats, suprarenal aortic infusions of 8-40 nmol/kg/hr CCPA decreased IMBF (15%) and other perfusion indices (22%-27%); in LS rats, IMBF increased 3% (insignificant) and other indices decreased 13%-24%. In LS rats, pretreatment with N-nitro-L-arginine methyl ester prevented the A1 receptor-mediated decrease in RBF and CBF but not OMBF; the response in IMBF was not altered. Pretreatment with indomethacin prevented the decreases in RBF, CBF, and OMBF and did not change the response of IMBF. Thus, within the cortex the vasoconstriction that follows A1 receptor activation results both from inhibition of NO synthesis and from stimulation of vasoconstrictor products of the COX pathway. In the outer medulla, the latter products seem exclusively responsible for CCPA-induced vasoconstriction. The observation that in LS rats IMBF was not affected by stimulation of adenosine A1 receptors suggests that limiting salt intake may help protect medullary perfusion against vasoconstrictor stimuli which have the potential to disturb long-term control of arterial pressure.

Furosemide-induced renal medullary hypoperfusion in the rat: role of tissue tonicity, prostaglandins and angiotensin II.

Furosemide (frusemide)-induced renal medullary hypoperfusion provides a model for studies of the dependence of local circulation on tissue tonicity. We examined the role of medullary prostaglandins (PG) and adenosine (Ado) as possible mediators of the response to furosemide. Furosemide was infused i.v. at 0.25 mg kg(-1) h(-1) in anaesthetized rats, untreated or treated with intramedullary indomethacin (Indo) or Ado. An integrated set-up was used to measure renal medullary laser-Doppler flux (MBF) and medullary ionic tonicity (electrical admittance, Y), and to infuse Indo and Ado directly into the medulla. The cortical flux was measured on kidney surface. The excretion of water, sodium and total solute was also determined. Intramedullary Indo (1 mg kg(-1) h(-1)) decreased MBF 18 +/- 5% and increased tissue Y 14 +/- 3% (both significant); the treatment abolished the post-furosemide decrease in MBF (-22% in untreated group) and enhanced slightly the increase in renal excretion. Intramedullary Ado (5 mg kg(-1) h(-1)) did not change baseline MBF or Y; the post-furosemide decreases in MBF (-22%) and Y, and the increase in renal excretion were preserved. We conclude that a decrease in intramedullary PG activity secondary to decreased medullary hypertonicity mediates the fall in medullary perfusion in response to furosemide; the hypoperfusion may help restore the initial tonicity. Together with the earlier evidence on the dependence of post-furosemide medullary hypoperfusion on angiotensin II, the study exposes its interaction with PG in the control of medullary circulation. Adenosine is not involved in medullary vascular responses to decreased tissue hypertonicity.

The influence of pH, temperature and buffers on the degradation kinetics of cefetamet pivoxil hydrochloride in aqueous solutions.

The first-order hydrolysis kinetics of cefetamet pivoxil (CP) were investigated as a function of pH, temperature and buffers. The degradation was followed by HPLC. Buffer catalysis was observed in acetate and phosphate buffers. The pH-rate profiles for hydrolysis of cefetamet pivoxil were obtained at 333, 343, 353 and 363K. The pH-rate expression was k(pH)=kH+aH+ + kH2OkOH-aOH-, where kH+ and kOH- are the second-order rate constants (mol(-1)ls(-1)) for hydrogen ion activity and for hydroxyl ion activity respectively, and kH2O is the pseudo-first-order rate constant (s(-1)) for spontaneous reaction under the influence of water. The pH-rate profile was characteristically U-shaped. Maximum stability was observed in the pH region from 3 to 5.

Renal medullary infusion of indomethacin and adenosine. Effects on local blood flow, tissue ion content and renal excretion.

Perfusion of the renal medulla and osmotic hypertonicity of its interstitium are the two important features of this zone which can influence body fluid homeostasis, especially arterial blood pressure. Separate manipulation of the two variables is best obtained with the intramedullary infusion of active agents. In this study, a set-up combining the possibility of infusion into the medulla with measurement of local blood flow (MBF, laser-Doppler flux) and extracellular ion concentration (tissue electrical admittance, Y) was used to determine effects of intramedullary indomethacin (Indo) and adenosine (Ado) in anaesthetized rats. Intramedullary Indo, 1 mg kg(-1 )h(-1), significantly increased tissue Y, by 12 +/- 3%, and significantly decreased MBF by 20 +/- 3%. There was also an unexplained increase of sodium excretion (U(Na)V) by 169 +/- 24% and of urine flow (V) by 62 +/- 6% (n = 10, both p < 0.03). Intramedullary Ado, 5 microg kg(-1) h(-1), did not alter Y, MBF or U(Na)V, whereas V increased 45 +/- 6% and urine osmolality decreased 25 +/- 4% (both changes significant). Elevation of medullary interstitial Ado to a level that did not alter MBF or U(Na)V induced a moderate defect of urine concentration that was not due to a decrease in ionic medullary hypertonicity.

Evaluation of stability of cefuroxime axetil in solid state.

The effect of temperature and relative atmospheric humidity on the stability of the crystalline form of cefuroxime axetil (CFA) in solid state was investigated. CFA is a mixture of diastereomers A and B. Changes in the concentration of the two diastereomers (A and B) of CFA were recorded by means of HPLC with UV detection. The degradation of diastereomers of CFA occurring at 0% relative humidity (RH) of the ambient air is a reversible first order reaction, while that occurring in humid air (RH>50%) is an autocatalytic first order reaction relative to substrate concentration. Although it has been found, that diastereomer B is the more stable isomer, humidity has a stronger effect on this very diastereomer.

The renal medullary interstitium: focus on osmotic hypertonicity.

1. There has been continued interest in the functional role of the renal medullary interstitium and intense research in this area has furnished new information regarding the extent, dynamics and mechanisms determining fluctuations in medullary osmotic hypertonicity. 2. Any change in the tonicity (interstitial solute concentration) indicates an imbalance of the rate of solute delivery to the interstitium (by tubular transport) and solute removal therefrom (by the microcirculation). It is often difficult to establish whether alteration of the delivery or removal triggered the change in medullary tissue tonicity. 3. Newer in vivo studies have confirmed earlier predictions and indirect evidence indicating that the rate of NaCl transport in the ascending limb of the loop of Henle is the major determinant of medullary ionic hypertonicity. 4. The hypothesis of a 'washout' of medullary solutes during increased medullary blood flow (MBF) has been re-evaluated. A novel experimental approach has provided direct evidence of a modest dissipation of medullary solutes with increasing MBF and a modest accumulation of solutes with decreasing MBF. 5. Increasing evidence is reviewed indicating that medullary tonicity is not only a regulated variable, but also that it may itself modulate the activity of multiple local endocrine and paracrine control systems and thereby affect local microcirculation and the function of medullary interstitial and tubular cells.