Involvement of nitric oxide in the regulation of regional hemodynamics in streptozotocin-diabetic rats.

In experimental and human diabetes mellitus, evidence for an impaired function of the vascular endothelium has been found and has been suggested to contribute to the development of vascular complications in this disease. The aim of the study was to evaluate possible regional hemodynamic in vivo differences between healthy and diabetic rats which would involve nitric oxide (NO). Central hemodynamics and regional blood flow (RBF) were studied using radioactive microspheres in early streptozotocin (STZ)-diabetic rats and compared to findings in healthy control animals. This method provides a possibility to study the total blood flow and vascular resistance (VR) in several different organs simultaneously. L-NAME iv induced widespread vasoconstriction to a similar extent in both groups. In the masseter muscle of both groups, acetylcholine 2 microg/kg per min, induced a RBF increase, which was abolished by pretreatment with L-NAME, suggesting NO as a mediator of vasodilation. In the heart muscle of both groups, acetylcholine alone was without effect while the combined infusion of acetylcholine and L-arginine induced an L-NAME-sensitive increase in RBF. The vasodilation induced by high-dose acetylcholine (10 microg/kg per min) in the kidney was more pronounced in the STZ-diabetic rats. The results indicate no reduction in basal vasodilating NO-tone in the circulation of early diabetic rats. The sensitivity to vasodilating effects of acetylcholine at the level of small resistance arterioles vary between tissues but was not impaired in the diabetic rats. In the heart muscle the availability of L-arginine was found to limit the vasodilatory effect of acetylcholine in both healthy and diabetic rats. In conclusion, the results indicate a normal action of NO in the investigated tissues of the early STZ-diabetic rat.

Regulation of uveal and retinal blood flow in STZ-diabetic and non-diabetic rats; involvement of nitric oxide.

PURPOSE: In experimental and human diabetes mellitus evidence for an impaired function of the vascular endothelial cells has been found. The purpose of the present experiments was to measure uveal and retinal blood flow and vascular resistance at an early stage of experimental diabetes mellitus and to evaluate the effects of acetylcholine and L-arginine in control and L-NAME-pretreated animals. METHOD: The radioactively labelled microsphere method was applied to normal Sprague-Dawley rats and rats with STZ-induced diabetes of three weeks duration. RESULTS: In the present study, similar blood flow and vascular resistance were observed in the uvea of normal and STZ-diabetic rats. Evidence for a basal vasodilating NO-tone was found both in the uvea and in the retina of both groups. In the normal rats as well as in the diabetic animals, acetylcholine induced choroidal vasodilation. Local blood flow increased from 54 +/- 17 to 142 +/- 32 mg x min(-1) in normal rats and from 57 +/- 18 to 112 +/- 23 mg x min(-1) in diabetic rats (P < 0.05 respectively). No hemodynamic changes were observed in the anterior uvea, demonstrating a difference in reactivity between these vascular beds. In animals pretreated with the NO-synthase inhibitor L-NAME, acetylcholine did not significantly affect local blood flow in the choroid, suggesting NO as a mediator of the vasodilation. CONCLUSION: The results indicate a normal action of NO in the ocular vascular beds at this stage of experimental diabetes mellitus in the rat.

Effects of endothelin receptor type A antagonism and nitric oxide synthase inhibition on cerebral blood flow in hypertensive rats.

Effects of the endothelin receptor type A antagonist BQ 123 and the NO synthase inhibitor L-NMMA on cerebral blood flow were studied in vivo in anaesthetized hypertensive (SHR) and normotensive (WKY) rats. The effects of acetylcholine following pre-treatment with these drugs were also studied with the microsphere method for blood flow determination in the cortex, thalamus, caudatus, pons, medulla, cerebellum and hypophysis. BQ 123 (1 mg kg-1) induced only minor effects on cerebral blood flow in both strains (n = 8), whereas L-NMMA (N = 8; 20 mg kg-1) reduced regional cerebral blood flow significantly in most regions (21-54%) in the hypertensive, but not in the normotensive rat. In normotensive rats pre-treated with BQ 123 intravenous administration of acetylcholine (2 micrograms kg-1 min-1) induced a widespread significant increase (20-50%) in cerebral blood flow despite a reduction of the mean arterial blood pressure, while no significant effects were seen in hypertensive animals. Intravenous infusion of acetylcholine in animals pre-treated with L-NMMA did not affect cerebral blood flow in most regions in either of the two rat strains. In conclusion, a vasodilatory response to acetylcholine was found following endothelin receptor A antagonism in the WKY rat only, suggesting a role for endothelin in the control of cerebral blood flow in this strain. Furthermore, a higher basal vasodilating nitric oxide-tone seems to be present in the hypertensive rat compared with the normotensive rat.

Effects of nitric oxide synthase inhibition and endothelin ETA receptor blockade on haemodynamics in hypertensive rats.

1. The objectives of the present study were to study regional differences in haemodynamics between spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats induced by the nitric oxide synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA) and the endothelin ETA receptor antagonist BQ 123 in vivo in tissues known to be important for blood pressure (BP) regulation (heart, kidney and skeletal muscle). Furthermore, the effect of acetylcholine (ACh) infusion (2 micrograms/kg per min) was examined after L-NMMA or BQ 123. The microsphere method was used for determinations of cardiac index (CI) and regional haemodynamics. 2. NG-Monomethyl-L-arginine (20 mg/kg) increased BP (26-48%; P < 0.01) and reduced CI in both rat strains. BQ 123 (1 mg/kg) reduced BP slightly (-4 to 11%; P < 0.05). 3. NG-Monomethyl-L-arginine significantly increased myocardial and skeletal muscle vascular resistance in SHR only; however, in the kidney, L-NMMA reduced blood flow and increased vascular resistance in both rat strains. 4. BQ 123 induced minor changes in regional haemodynamics that were not significantly different between the two strains. 5. Acetylcholine following BQ 123 induced an increase in myocardial blood flow in WKY rats, but decreased blood flow in SHR. Acetylcholine following L-NMMA reduced myocardial blood flow in both strains. 6. Acetylcholine following BQ 123 induced renal vasodilation in WKY rats but, following L-NMMA, ACh did not induce renal vasodilation in either rat strain. In contrast, L-NMMA did not abolish the vasodilation of acetylcholine in skeletal muscle in WKY rats. 7. In conclusion, the contribution of nitric oxide to basal vessel tone was not impaired in the heart, skeletal muscle and kidney in SHR. Antagonism of ETA receptors caused similar haemodynamic responses in both rat strains in these organs. Furthermore, NOS inhibition, but not ETA blockade, blunted the expected ACh-induced vasodilation in the heart and kidney in WKY rats, but not in skeletal muscle in both strains.

Regional haemodynamic differences between normotensive and spontaneously hypertensive rats--a microsphere study.

The objective of the present study was to compare systemic and regional haemodynamics in a large series of spontaneously hypertensive rats (SHR, n=32) with normotensive Wistar-Kyoto rats (WKY, n=26) at the age of 12-16 weeks. All rats were anaesthetized with thiobutabarbital and the radioactively labelled microsphere method was used to evaluate regional blood flow with special emphasis on different cerebral areas. The high blood pressure in the SHR was mainly due to elevated total peripheral resistance, which was 90% higher in the SHR compared to the WKY. Furthermore, heart rate was 25% (p<0.001) higher, but the cardiac index was lower by 20% (p<0.01) in the SHR. Blood flow was significantly lower and vascular resistance higher in several organs such as the kidneys, other visceral organs, skeletal muscle and skin of the SHR compared to the WKY. On the contrary, blood flow in the myocardium was augmented by 40% (p<0.01) in the SHR. Blood flow was 20-50% higher in the cerebral cortex, thalamus and caudatus (p<0.05-0.001), but attenuated in the hypophysis of the SHR. In the pons, medulla and cerebellum, blood flow was similar in the two strains. In this large microsphere study, the basal cardiac index was lower in the SHR already at this relatively early stage of established hypertension. Despite this, increased blood flow in the above mentioned cerebral regions was found in the SHR compared to the WKY.

Endothelium-dependent vasodilation in the uvea of hypertensive and normotensive rats.

PURPOSE: The effects of endothelium-related substances such as acetylcholine, a stimulator of endogenous NO-production, the NO-synthesis inhibitor L-NMMA, the exogenous NO-donor sodium nitroprusside and the endothelin (ET)A-receptor antagonist BQ123, on uveal blood flow were investigated in normotensive and hypertensive SHR rats. METHOD: The radioactively-labelled microsphere method was applied for the measurement of regional blood flow in the uvea. RESULTS: Under resting conditions, local blood flow was lower in the hypertensive animals. The increase in choroidal blood flow (145 +/- 50%; P < 0.01) and reduction in vascular resistance (-58 +/- 7%; P < 0.01) observed in the WKY after i.v. infusion of acetylcholine, 2 micrograms x kg bw-1 x min-1, were significantly less pronounced in animals pretreated with L-NMMA, indicating local formation of NO as a vasodilator mechanism. In contrast, acetylcholine did not induce significant vasodilation in the choroid of SHR rats. In the anterior uvea of both strains, acetylcholine did not affect local blood flow. L-NMMA, 20 mg x kg bw-1, alone reduced blood flow in the entire uvea of both strains. Intravenous injection of BQ123, 1 mg x kg bw-1, did not affect regional blood flow in the uvea of WKY or SHR animals. Infusion of acetylcholine following ETA-receptor blockade induced vasodilation in both the choroid and anterior uvea in the WKY but not in the SHR. CONCLUSIONS: Acetylcholine-stimulated NO-mediated vasodilation, but not basal NO-formation, was impaired in the choroid of the SHR. Furthermore, an interaction between vasoconstricting ET and acetylcholine was found in the anterior uvea of normotensive but not hypertensive rats.

Effects of acetylcholine and nitroprusside on systemic and regional hemodynamics in hypertensive rats.

This study was performed to investigate the in vivo effects of acetylcholine, a stimulator of endogenous NO production, and nitroprusside, an exogenous NO-donor, on hemodynamics in the normotensive (WKY) and the hypertensive (SHR) rat. Anesthetized rats were given microspheres for the measurement of cardiac index (CI), total vascular resistance (TPRI), regional blood flow and vascular resistance. Infusion of acetylcholine (2 microg/kg/min) caused a marked decrease in TPRI by (-35+/-5%, +/-SEM) in the WKY (n=8), whereas in the SHR (n=8) a less pronounced reduction was seen (-14+/-3%, p<0.01 between groups). CI increased by 27+/-9% in the WKY, but was unaltered in the SHR. Blood pressure decreased similarly (17-20%). Acetylcholine significantly increased blood flow by about 40% in the kidneys and the heart in the WKY, but had no significant effect in the SHR. Other tissues, such as skeletal muscle and cerebral tissues, showed no major changes. Infusion of nitroprusside (1 microg/kg/min) reduced blood pressure by 5 to 10% in the strains. The regional effects of nitroprusside did not differ between the strains. In conclusion, the acetylcholine-induced vasodilation in the kidney and the heart was attenuated in the SHR compared to the WKY. These findings might suggest a difference in the endothelial response between the SHR and the WKY in some, but not in all, tissues.

Vascular effects of endothelin-1 in the cat; modification by indomethacin and L-NAME.

Intravenous infusion of endothelin-1 (ET-1) in the cat, 60 pmol x kg body wt-1 x min-1 for 5 min, induced an increase in mean arterial blood pressure (MAP) of 41.3 +/- 4.8 mmHg (n = 6; P < 0.001). Blood flow, as determined with radioactive microspheres, was reduced in many tissues. Reductions by 70-80% were observed in the choroid plexus, pineal and pituitary glands. Total cerebral blood flow was reduced by 18-23%. Pre-treatment with indomethacin or a combination of indomethacin and L-NAME caused vasoconstriction in many tissues and modified the responses to ET-1 in a variable way, suggesting that normally, ET-1 tends to release arachidonic acid metabolites and nitric oxide with great variations between different tissues. Intracerebroventricular infusion (i.c.v.) of ET-1, 10 pmol x kg body wt-1 x min-1, caused an increase in MAP of 79 +/- 11 mmHg (n = 6; P < 0.001). Regional blood flow in the medulla oblongata, medulla spinalis, choroid plexus, pineal and pituitary glands was reduced by 60-80%. Heart rate, cardiac output and coronary blood flow were significantly increased after 30 min i.c.v. infusion, indicating an activation of the heart, most probably as part of a central ischaemic response. Our results indicate that in many tissues the vasoconstrictive effect of ET-1 is influenced by indomethacin- and L-NAME-sensitive vasodilator mechanisms that are activated by the peptide. In the CNS, there may be marked effects on regional blood flow after i.c.v. infusion.

Ocular effects of endothelin-1 in the cat.

The effects of intravitreal and intracameral administration of endothelin-1 were studied in the anaesthetized cat. Intravitreal injection of 0.4 nmol endothelin-1 induced a 34 +/- 10% (n = 5; P less than 0.05) reduction in retinal blood flow in the experimental eye compared to the control eye 80 minutes after the injection. Blood flow in the ciliary body, iris and choroid remained unaffected by the injection of endothelin-1. Intracameral administration of endothelin-1 at pmol doses caused a reduction in pupil size, an increase in the aqueous humor protein concentration (indicating a break-down of the blood-aqueous barrier) and an increase in the concentration of prostaglandin E2 in the aqueous humor. The effect of endothelin-1 on pupil size was abolished by indomethacin pretreatment, indicating that this effect was mediated by arachidonic acid metabolites.

Effects of cervical sympathetic nerve stimulation and neuropeptide Y (NPY) on cranial blood flow in the cat.

Effects of cervical sympathetic nerve stimulation (SNS) at 10 Hz and intravenous infusion of neuropeptide Y (NPY), 10 and 100 pmol x kg body wt-1 x min-1 for 5 min, on regional blood flow in the cat were investigated with radioactive microspheres. Sympathetic nerve stimulation caused significant reductions in blood flows in the facial tissues including the eye. Alpha-adrenoceptor blockade with phenoxybenzamine and combined beta- and alpha-adrenoceptor blockade with propranolol and phenoxybenzamine abolished the effects of sympathetic nerve stimulation in most facial tissues except in the tongue, upper eyelid and masseter muscle. In most cranial tissues, neuropeptide Y reduced regional blood flow and increased vascular resistance. No effect of neuropeptide Y on vascular resistance was observed in the choroid. In the present study, evidence for a non-adrenergic component in sympathetic vasoconstriction was found in the tongue, upper eyelid and masseter muscle but not in the majority of feline facial tissues. Neuropeptide Y was a potent vasoconstrictor in many cranial tissues, while in parts of the uvea, the effects of neuropeptide Y were less pronounced.

Effects of endothelins (ET-1, ET-2 and ET-3) in the rabbit eye; role of prostaglandins.

The effects of intracameral injection of three endothelin isopeptides (ET-1, ET-2 and ET-3) were studied in the rabbit eye. All three peptides at pmol doses caused a dose-dependent rise in intraocular pressure (IOP) and an increase in protein concentration in the aqueous humor, indicating breakdown of the blood-aqueous barrier. Mean arterial blood pressure was not affected. ET-1 was more effective to increase IOP than were ET-2 and ET-3 whose effects seemed to be equal. ET-1 also caused vasodilation in the anterior uvea. The effects of ET-1, ET-2 and ET-3 on the IOP and blood-aqueous barrier, as well as the effects of ET-1 on regional blood flow in the eye, were abolished by pretreatment with indomethacin. Injection of 4 pmol ET-1 into the anterior chamber caused an increase in the concentration of prostaglandin E2 (PGE2) in the aqueous humor, most probably as a result of increased production of PGE2 in the anterior uvea. The results indicate that the effects of ET-1, ET-2 and ET-3 in the rabbit eye are to a large extent mediated by arachidonic acid metabolites.

Non-adrenergic sympathetic vasoconstriction in the eye and some other facial tissues in the rabbit.

The effects of unilateral sympathetic nerve stimulation (SNS) on regional blood flow in the rabbit were studied with radioactive microspheres. SNS at 10 or 4 Hz caused an approximately 60% reduction in choroidal blood flow, which was partly resistant to alpha-adrenoceptor blockade with phenoxybenzamine. The vasoconstriction evoked by SNS at 2 Hz was completely abolished by alpha-adrenoceptor blockade. A similar response was seen in the iris, ciliary body, masseter muscle and lacrimal gland. In the harderian gland, however, SNS (2 Hz) after alpha-adrenoceptor blockade caused a significant reduction in blood flow. In the salivary glands, combined beta- and alpha-adrenoceptor blockade with propranolol and phenoxybenzamine revealed a slight non-adrenergic vasoconstriction during SNS at 10 Hz; however, the blood flow was significantly increased during SNS at 4 and 2 Hz following alpha-adrenoceptor blockade. These results indicate that there is a frequency-dependent, non-adrenergic component in the sympathetic vasoconstriction of the eye and several facial tissues. In the salivary glands, beta-adrenoceptor-mediated vasodilatation tends to mask a non-adrenergic vasoconstriction.