Jean-Jacques Serrano

Effects of vanadyl derivatives on animal models of diabetes

Vanadium is a transition metal of the Vb group found in living organisms at concentrations varying from the pM to the/xM range. It is a growth factor for various plants and is considered as essential in some animal species, such as chicken or rat, for which a complete vanadium deprivation in food induces impairment of growth and reproductive performances [1]. In the human species, its essentiality has not been established. Vanadium possesses a number of biochemical properties in vitro [1]. In particular, it is a potent Na+/K + ATPase inhibitor at concentrations close to the nM range, which has suggested that it might play the role as a physiological regulator of the enzyme. However, this inhibition is specific of the 5+ form (vanadate) of the metal, which is mostly found extracellularly in living organisms. Intracellularly, vanadium is reduced into the 4+ form (vanadyl), and the intracellular 5+ form concentration is probably too low to play a significant role in physiological conditions [2]. Since 1979, vanadium in the form of sodium vanadate has been shown to possess insulin-like properties on various cellular models, such as adipocytes, muscular cells or isolated hepatocytes [3-8]. In particular, vanadate stimulates glucose transport, inhibits glycogenolysis, and inhibits lipolysis. Although this insulin-like activity was obtained in vitro using relatively high concentrations (from the/xM to the mM range), a number of studies have been conducted to study its mechanism. Various hypotheses are still being debated, such as (a) inhibition of phosphotyrosyl-protein phosphatases [9], (b) direct induction of tyrosine phosphorylation [10-12] although this hypothesis was recently challenged [13], (c) direct activation of glucose transport through activation of the GLU4 transporter gene [14] or translocation of the transporter from the cellular organelles to the plasma membrane [15], (d) increase of intracellular calcium secondary to Ca-Mg-ATPase inhibition [16], or (e) stimulation of NADH oxydase and H202 formation [1], a mechanism supported by the mutual potentiation of vanadate and H202 demonstrated in vitro [17[. The interest for the use of vanadium in the diabetic state was brought about with the first demonstration by Heyliger and coworkers of its antidiabetic activity in the streptozotocin (STZ)-treated rat [18]. Administered in drinking water at a concentration of 0.8 mg/ml, sodium orthovanadate was able to control hyperglycemia and to prevent the impairment of cardiac function. An interesting result was the lack of increase of plasma in-

Impairment of contractile response to carbachol and muscarinic receptor coupling in gastric antral smooth muscle cells isolated from diabetic streptozotocin-treated rats and db/db mice

This work explored the role of the cholinergic pathway, assessed at a post-synaptic level by the use of isolated smooth muscle cells, in the impairment of antral motility associated with diabetic gastroparesis.Contractile response to carbachol — but not to erythyromycin, a motilin receptor agonist — was abolished in antral smooth muscle cells isolated from (i) rats previously rendered diabetic by a single i.v. dose of streptozotocin (STZ, 60 mg/kg) and (ii) db/db spontaneously diabetic mice. Insulin treatment of STZ-rats was able to prevent the impairment of the carbachol contractile response, but not to reverse it once established. In STZ-rats, impairment of contractile response was not associated with a change in density of [3H]-N-methyl-scopolamine ([3H]-NMS) binding sites (≈ 1.5 fmol/mg protein). Displacement curve of the [3H]-NMS binding by carbachol was shifted to the right in diabetic rats as compared to controls. The addition of GTP-γ-S induced a shift to the right of the displacement curve in control but not in diabetic animals.These results strongly suggest that diabetes is associated with an early and specific alteration of the muscarinic control of contraction of antral smooth muscles at a post-synaptic level, associated with an alteration of the GTP-binding proteins coupled to muscarinic receptors.

Circadian and circannual variation of the carregeenin inflammatory effect in rat

The circadian variation of edema produced by carrageenin (carr.) administration into plantar tissue was studied in rats kept under a 12 light - 12 dark regimen. Three doses were used (125, 250 and 500 micrograms per rat) injected at different time (02.00, 08,00, 14.00 and 20.00 h). With the high doses, the level of edema for the four hour period after carr. administration was similar whatever the hour of injection. In contrast, with the lower dose (125 micrograms) a circadian rhythm in the intensity of the edema produced was observed, showing a maximum of susceptibility during the light span. Repetitive experiments performed at different periods of the year validated this finding. Comparing mean mesors, analysis of this data showed two distinct levels of inflammation, with the lower level observed in autumn and winter indicating evidence for a circannual variability.

Estimation of pharmacokinetic parameters of sodium tungstate after multiple-dose during preclinical studies in beagle dogs

In this paper, an empirical Bayes methodology was used to determine the pharmacokinetic profile of sodium tungstate in beagle dogs after multiple oral dosing using the P-PHARM computer program. The population estimation algorithm used in P-PHARM is an EM-type procedure. Sodium tungstate was administered orally, three times a day, (i) for 11 days (21 and 42 mg/kg per day) to 18 dogs (nine males and nine females) and (ii) for 13 weeks (15, 30 and 60 mg/kg per day) to 28 dogs (14 males, 14 females). Six other dogs received the compound intravenously (25 and 50 mg/kg). Plasma concentration profiles versus time were compatible with a two-compartment model and first-order kinetics. After oral administration, F (0.61+/-0.086 vs. 0.48+/-0.093), and normalized (to a 7-mg/kg dose of sodium tungstate) AUC (54+/-8.4 vs. 41.2+/-8.5 mg/l x h), C(max) (10.6+/-0.49 vs. 8.5+/-0.57 microg/ml) and C(min) (3.04+/-0.23 vs. 2.04+/-0.22 microg/ml), were higher in male than in female dogs. However, the introduction of the gender in the final model did not contribute statistically to an improvement of the fit of the population pharmacokinetic model. In males, t(1/2) elimination averaged 3.1+/-0.56 vs. 2.6+/-0.18 h in females. The duration of treatment did not modify statistically the pharmacokinetic parameters. After repeated multiple oral administration of 15-60 mg/kg per day of sodium tungstate, tungsten plasma concentrations increased in proportion to dose. No dose-dependent changes in pharmacokinetic parameters occurred.

Vanadium pharmacokinetics and oral bioavailability upon single-dose administration of vanadyl sulfate to rats.

Vanadium pharmacokinetic parameters and oral bioavailability were determined after administration of vanadyl sulfate, an antidiabetic agent, to male Wistar rats. An optimal sampling design was used over a 21‐day period; vanadium was measured in blood by atomic absorption spectrophotometry (AAS). After i.v. bolus injection (3.025 mg V/kg body weight), a three‐compartment model was fitted to the data. Mean (± SD) half‐lives were 0.90 ± 0.56 hours, 24.8 ± 14.5 h and 201 ± 74 h, respectively, for the three phases observed. Vanadium clearance averaged 37.6 ± 15.8 mL/h. Initial volume of distribution was 2.43 ± 1.22 L/kg whereas total volume of distribution was 25.4 ± 3.9 L/kg; these values largely exceeded body weight (i.e. 300 g), in agreement with a great uptake and retention of vanadium in tissues. After oral gavage administration (15.12 and 7.56 mg V/kg body weight), vanadium disposition was best described by a three‐compartment model, with absorption appearing to occur by a zero‐order rate. This process lasted 10.3 ± 1.3 h and 10.9 ± 1.1 h for the two dosage levels, respectively. Half‐lives corresponding to the terminal log‐linear part of the curve were 173.5 ± 1.6 h and 172 ± 6 h (Bayesian estimates). No dose‐dependency was observed for any of the parameters determined. Absolute bioavailabilities, with reference to the i.v. administration, were 12.5% and 16.8% when determined from AUCmod. Bioavailability appeared to be higher than generally stated in the literature.

Post-natal evolution of rat cardiac beta-adrenoceptors

Cardiac beta-adrenoceptors (beta AR) were studied using membranes prepared at birth (day 0) and at days 7, 10, 15, 21, 30, 45 and 60. Saturation experiments using the antagonist ligand (125I)-iodocyanopindolol (ICYP) allowed the determination of beta AR number (Bmax) and ICYP dissociation constant (Kd), while (-)isoproterenol competition curves of ICYP binding, performed in the absence or presence of Gpp(NH)p (10(-4) M), were used to measure the relative proportions of high and low affinity states of the beta AR for the agonist and to assess the ability of beta AR to couple with the GTP-binding protein. Rat cardiac beta AR evolved at 3 distinct periods: during the first period (days 0-10), the receptor density and ICYP Kd were half that of adults, and beta AR were present only in an homogeneous high affinity state. The second period (days 15-21) was characterized by a progressive increase in beta AR number and ICYP Kd, while analysis of (-)isoproterenol competition curves indicated that beta AR were poorly coupled to the GTP-binding protein. In the third period (days 30-60), ICYP Bmax and Kd were respectively 53.9 +/- 1.2 fmoles/mg protein and 106.4 +/- 2.9 pM, while analysis of (-)isoproterenol competition curves showed the existence of high and low affinity binding states in equal proportions in the absence of Gpp(NH)p, and of one homologous low affinity state of the receptor in its presence. These data indicate that beta AR follow a postnatal evolution marked by an increase in beta AR density concomitant with a decrease in affinity toward the antagonist ligand ICYP, accompanied by the progressive appearance of a poorly-coupled beta AR. However, the number of efficiently coupled receptors was found to be similar in adult and newborn rats.

Vasopressin V2 (SR121463A) and V1a (SR49059) receptor antagonists both inhibit desmopressin vasorelaxing activity

Although [Arg(8)]vasopressin is a potent vasoconstrictor, it possesses vasorelaxant properties manifested either after vasopressin V1 receptor blockade or directly in some vascular beds. The nature of the receptor involved in the vasorelaxant effect of [deamino-Cys(1) D-Arg(8)]vasopressin (desmopressin), a vasopressin V2 receptor agonist, was studied on rat precontracted aortic rings by the use of highly selective new non-peptide vasopressin receptor antagonists. The present study demonstrates for the first time that desmopressin relaxant effect is antagonized by the vasopressin V2 receptor antagonist SR121463A, but also by the vasopressin V1A receptor antagonist SR49059, suggesting that desmopressin-induced relaxation is mediated by a receptor subtype sharing both V1A and V2 pharmacological profiles.

Cardiac adenylate cyclase activity in streptozotocin-treated rats after 4 months of diabetes: impairment of epinephrine and glucagon stimulation

Adenylate cyclase (AC) activities of cardiac membranes prepared (a) from rats that had been made diabetic 4 months previously by a single i.v. injection (50 mg/kg) of streptozotocin (STZ), and (b) from diabetic rats which had been treated during the same period by a daily dose of long-acting insulin (2-4 U/animal), were compared with the AC activity of cardiac membranes prepared from age-matched control animals. Basal (Mg++-dependent) and Mn++ (7 mM)-dependent activities, as well as Gpp(NH)p (3 X 10(-7) M) stimulation and Ca++ (pCa = 3.9) inhibition of cardiac AC were not significantly different in the three groups. At the EC50 concentration epinephrine (5 X 10(-7) M) stimulation of AC was reduced in diabetic animals but no change was observed at higher concentrations (10(-4) M). Glucagon stimulation was impaired at both the EC50 concentration (10(-7) M) and at higher concentrations (10(-5) M). Insulin treatment of the diabetic animals partially prevented the impairment of hormone stimulation. These results confirm observations that alterations of cardiac AC activity in STZ-treated rats are indeed due to diabetes and not to STZ-toxicity and suggest that AC-coupled receptors are altered either by an diabetes-induced alteration of cardiac sarcolemma.

Vanadyl sulphate differently influences insulin response to glucose in isolated pancreas of normal rats after in vivo or in vitro exposure

The effect of the antidiabetic agent vanadyl sulphate (VOSO4) on the endocrine pancreas function of normal rats was studied using the isolated pancreas preparation. A short-term (8 days) i.p. treatment (15 mg/kg per day) resulted in attenuation of high glucose-stimulated insulin release, at day 9 but also at days 19, i.e., after full recovery of appetite and weight, while blood and pancreas vanadium concentrations were still elevated. Six months of oral VOSO4 treatment (0.75 mg/ml in drinking water) resulted in elevated vanadium concentrations while glucose-stimulated insulin release was attenuated as compared to pair-fed animals. Conversely, when directly perfused in pancreas, VOSO4 potentiated glucose-stimulated insulin release. These apparently opposite effects may be related to the ability of VOSO4 to exert both peripheral insulinomimetic effects-leading to chronic reduction in insulin demand-, and a direct pancreatic insulinotropic activity.

Impairment of contractility associated with muscarinic supersensitivity in trachea isolated from diabetic rats: lack of correlation with ultrastructural changes or quinuclidinyl benzylate binding to lung membranes

Evolution of cholinergic response of rat isolated trachea was determined after various durations of diabetes (17, 40, 90, 150 and 210 days). Long-term diabetes was associated with both impairment of contractility and supersensitivity to cholinergic stimulation. However, the mechanism of these alterations remains to be determined, as response to field stimulation was not specifically altered while electron microscopy studies could not detect any significant change in the aspect of nerves, smooth muscle or epithelium. As well, binding studies of lung cholinergic receptors using the antagonist ligand [3H]-quinuclidinyl benzylate and the agonist carbachol did not detect any change in diabetic animals.