title: In vivo and in vitro evidence of altered nitric oxide metabolism in the spontaneously diabetic, insulin-dependent BB/Edinburgh rat.
abstract: 1. Altered vasoreactivity may contribute significantly to the pathogenesis of diabetic vascular complications. This study investigated the effect of (a) insulin-related diabetes, and (b) chronic in vivo administration of N(omega)-nitro-L-arginine ester (L-NAME), a nitric oxide (NO) synthase inhibitor, on mean arterial pressure and in vitro vascular reactivity to noradrenaline in mesenteric arterial bed preparations from spontaneously diabetic, insulin-dependent and treated BB rats, the best animal model of insulin-dependent mellitus (IDDM) currently available. Four groups of animals from the Edinburgh colony (BB/E) of spontaneous diabetic BB rats were studied: age-matched (mean +/- s.e. mean = 156 +/- 2d) non-diabetic (glycated haemoglobin = 3.8 +/- 0.1%) and insulin-treated diabetic (glycated haemoglobin = 6.2 +/- 0.5%; duration of diabetes = 56 +/- 4 d) groups were either L-NAME treated (oral dose = 27 +/- 1 mg kg-1 d-1; duration of treatment from 30 until 153 days of age) or untreated. Although our diabetic BB/E rats do not achieve overall normoglycaemia, individual adjustment of the daily insulin dose administered to every diabetic rat achieves better glycaemic control than previous groups studying altered vascular reactivity and endothelial dysfunction in this animal model of diabetes. 2. Mean arterial pressure (measured directly via indwelling carotid arterial cannulae) was not significantly different between non-diabetic (116 +/- 3 mmHg; n = 10) and diabetic (122 +/- 2 mmHg; n = 12) BB/E rats. L-NAME treatment significantly (P < 0.001) increased mean arterial pressure in both groups (165 +/- 6 mmHg; n = 9 and 142 +/- 4 mmHg; n = 6 respectively) but the degree of hypertension observed in L-NAME-treated diabetic rats was significantly (P < 0.01) attenuated compared to non-diabetic rats treated with L-NAME. 3. Mesenteric arterial bed preparations were cannulated under anesthesia, excised and intralumenally perfused ex vivo with noradrenaline (0.2-20 microM). Basal perfusion pressures were not significantly different in mesentery preparations from non-diabetic (27.0 +/- 2.6 mmHg) and diabetic (27.1 +/- 3.2 mmHg) BB/E rats. There was no significant difference in maximal response above basal perfusion pressure (MAX) or pEC50, defined as the negative log of the agonist concentration required to give 50% of the maximal response above basal perfusion pressure, to noradrenaline in untreated non-diabetic (166 +/- 7 mmHg and 5.74 +/- 0.05 respectively) and diabetic (170 +/- 11 mmHg and 5.59 +/- 0.05) BB/E rats. 4. In vivo treatment of non-diabetic and diabetic BB/E rats with L-NAME had no significant effect on basal perfusion pressure (25.9 +/- 4.3 mmHg and 28.5 +/- 3.9 mmHg respectively). L-NAME treatment in vivo increased (P < 0.001) MAX to noradrenaline of non-diabetic rats (224 +/- 8 mmHg) but did not affect the value for diabetic rats (178 +/- 14 mmHg). L-NAME treatment did not alter after the pEC50 values in either group (5.71 +/- 0.05 and 5.65 +/- 0.05). 5. Consistent with previous studies using vascular preparations from spontaneously diabetic BB rats, mesentery preparations from diabetic BB/E rats (n = 12) exhibited a significantly reduced vasodilator response to acetylcholine (F value = 4.4, P < 0.05) across the concentration range studied compared to non-diabetic BB/E rats (n = 12) although there was no significant difference in maximal relaxation (diabetic 53.1 +/- 4.3% vs non-diabetic 55.7 +/- 5.5%) or pEC50, (diabetic 6.92 +/- 0.25 vs non-diabetic 7.49 +/- 0.22). There was no significant (F value = 0.8, P > 0.1) difference in the response to GTN between preparations from non-diabetic and diabetic rats (maximal relaxation: 49.6 +/- 3.7% vs 48.5 +/- 4.3%; pEC50: 7.84 +/- 0.12 vs 7.89 +/- 0.22 respectively). 6. In conclusion, vascular responsiveness to noradrenaline is not impaired in spontaneously diabetic BB/E rats with significantly better glycaemic control than those used in previous studies. However, following chronic L-NAME treatment, diabetic BB/E rats exhibit attenuated hypertension and an absence of enhanced vascular responsiveness to noradrenaline in vitro compared to similarly treated non-diabetic rats. These results, together with the significantly impaired endothelium-dependent vasodilatation and unchanged endothelium-independent vasodilatation in vitro of preparations from diabetic BB/E rats, are consistent with the hypothesis that functional changes in the synthesis and metabolism of NO (rather than altered vascular responsiveness to NO) occur in diabetes. Our results indicate that good glycaemic control alone is insufficient to prevent these abnormalities in NO availability and further studies to characterize the origin of these changes are necessary.
authors.affiliation: Metabolic Unit, Western General Hospitals NHS Trust, Edinburgh.
authors.forename: R M
authors.initials: RM
authors.lastname: Lindsay
authors.forename: R S
authors.initials: RS
authors.lastname: Peet
authors.forename: G S
authors.initials: GS
authors.lastname: Wilkie
authors.forename: S P
authors.initials: SP
authors.lastname: Rossiter
authors.forename: W
authors.initials: W
authors.lastname: Smith
authors.forename: J D
authors.initials: JD
authors.lastname: Baird
authors.forename: B C
authors.initials: BC
authors.lastname: Williams
mesh_terms: D000818:Animals; D001786:Blood Glucose / Q000378:metabolism; D001794:Blood Pressure / Q000187:drug effects; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D007004:Hypoglycemic Agents / Q000494:pharmacology; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology; D008297:Male; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism* / Q000502:physiology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D009638:Norepinephrine / Q000502:physiology; D051381:Rats; D011919:Rats, Inbred Strains; D013152:Splanchnic Circulation / Q000187:drug effects; D014661:Vasoconstriction / Q000187:drug effects; D014664:Vasodilation / Q000187:drug effects
chemical_list: D001786:Blood Glucose; D004791:Enzyme Inhibitors; D007004:Hypoglycemic Agents; D007328:Insulin; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D019331:NG-Nitroarginine Methyl Ester; D009638:Norepinephrine