Rat pial microvascular responses to melatonin during bilateral common carotid artery occlusion and reperfusion

The present study assessed the in vivo rat pial microvascular responses induced by melatonin during brain hypoperfusion and reperfusion (RE) injury. Pial microcirculation of male Wistar rats was visualized by fluorescence microscopy through a closed cranial window. Hypoperfusion was induced by bilateral common carotid artery occlusion (BCCAO, 30 min); thereafter, pial microcirculation was observed for 60 min. Arteriolar diameter, permeability increase, leukocyte adhesion to venular walls, perfused capillary length (PCL), and capillary red blood cell velocity (V(RBC) ) were investigated by computerized methods. Melatonin (0.5, 1, 2 mg/kg b.w.) was intravenously administered 10 min before BCCAO and at the beginning of RE. Pial arterioles were classified in five orders according to diameter, length, and branchings. In control group, BCCAO caused decrease in order 2 arteriole diameter (by 17.5 ± 3.0% of baseline) that was reduced by 11.8 ± 1.2% of baseline at the end of RE, accompanied by marked leakage and leukocyte adhesion. PCL and capillary V(RBC) decreased. At the end of BCCAO, melatonin highest dosage caused order 2 arteriole diameter reduction by 4.6 ± 2.0% of baseline. At RE, melatonin at the lower dosages caused different arteriolar responses. The highest dosage caused dilation in order 2 arteriole by 8.0 ± 1.5% of baseline, preventing leakage and leukocyte adhesion, while PCL and V(RBC) increased. Luzindole (4 mg/kg b.w.) prior to melatonin caused order 2 arteriole constriction by 12.0 ± 1.5% of baseline at RE, while leakage, leukocyte adhesion, PCL and V(RBC) were not affected. Prazosin (1 mg/kg b.w.) prior to melatonin did not significantly change melatonin's effects. In conclusion, melatonin caused different responses during hypoperfusion and RE, modulating pial arteriolar tone likely by MT1 and MT2 melatonin receptors while preventing blood-brain barrier changes through its free radical scavenging action

Protective Effects of Quercetin on Rat Pial Microvascular Changes during Transient Bilateral Common Carotid Artery Occlusion and Reperfusion

The aim of this study was to assess the in vivo effects of quercetin on pial microvascular responses during transient bilateral common carotid artery occlusion (BCCAO) and reperfusion. Rat pial microcirculation was visualized by fluorescence microscopy through a closed cranial window. Pial arterioles were classified in five orders of branchings. Capillaries were assigned order 0, the smallest arterioles order 1, and the largest ones order 5. In ischemic rats, 30 min BCCAO and 60 min reperfusion caused arteriolar diameter decrease (by 14.5 ± 3.3% of baseline in order 2), microvascular leakage [0.47 ± 0.04, normalized gray levels (NGL)], leukocyte adhesion in venules (9 ± 2/100 μm venular length, v.l./30 s), and reduction of capillary perfusion (by 40 ± 7% of baseline). Moreover, at the end of BCCAO and reperfusion there was a significant increase in reactive oxygen species (ROS) formation when compared with baseline. Quercetin highest dose determined dilation in all arteriolar orders (by 40 ± 4% of baseline in order 2) and prevented microvascular permeability (0.15 ± 0.02 NGL), leukocyte adhesion (3 ± 1/100 μm v.l./30 s) as well as ROS formation, while capillary perfusion was protected. Inhibition of endothelial nitric oxide synthase (NOS) prior to quercetin reduced arteriolar dilation (order 2 diameter increase by 10.3 ± 2.5% of baseline) and caused permeability increase (0.29 ± 0.03 NGL); inhibition of neuronal NOS or inducible NOS did not affect quercetin-induced effects. Inhibition of guanylyl cyclase prior to quercetin reversed the quercetin’s effects on pial arteriolar diameter and leakage. In conclusion, quercetin was able to protect pial microcirculation from ischemia–reperfusion damage inducing arteriolar dilation likely by nitric oxide release. Moreover, quercetin scavenger activity blunted ROS formation preserving the blood–brain barrier integrity

The Hypotensive and bradycardic effects of mouth opening: evidence in an animal model.

Objective Previous studies in normotensive anesthetized rats (Lapi Arch.Ital.Biol 151:11-23,2013) showed that peripheral stimulation of the trigeminal nerve induced by submaximal mouth opening (mandibular extension, ME) caused prolonged (at least 80min) bradycardia, hypotension and cerebral hemodynamic changes (pial arterioles showed a characteristic response pattern consisting in a significant constriction during ME followed by a dilatation for the entire remaining observation time). Design and method In this study we assessed the in vivo effects of ME on HR, MABP and pial microcirculation in hypertensive rats. Experiments were performed in male Wistar rats weighing 250-300g (n = 8). Hypertension was induced by intraperitoneal daily injection of dexamethasone (0.03mg/kg/day) for 10 days. ME was obtained by inserting an ad hoc developed retractor between the dental arches. HR and MABP were recorded by ECG and a catheter placed in the left femoral artery and measured by a computer-assisted system. Pial arterioles were observed through a closed cranial window implanted above the left parietal cortex and visualized by an in vivo fluorescence microscopy technique to assess vessel diameter changes before (baseline), during 10min ME and thereafter until 160min. Arteriolar diameters were measured with a computer-assisted method (MIP Image program, frame by frame). Results In sham-treated (no ME) hypertensive rats (n=3) HR, MABP and pial microcirculation did not change during whole observation period. Hypertensive rats subjected to ME (n=5) showed a significant decrease of HR and MABP. HR declined by 42bpm, (p<0.01) starting from 60 min after ME up to 160min, while MABP by 18mmHg (p<0.05) starting from 20min after ME up to 100min, compared with baseline. Pial arterioles exhibited a biphasic response: the arteriolar diameter decreased by 2.94&#956;m (p<0.05) during ME, afterwards it significantly increased by 3.46&#956;m (p<0.01) starting from 20min after ME; this vasodilatation lasted for the whole observation period. Conclusions Our results suggest that ME is able to exert profound and prolonged regulatory effects on systemic arterial blood pressure and pial arteriolar tone in hypertensive rats

Effects of Propionyl-L-Carnitine on Ischemia–Reperfusion Injury in Hamster Cheek Pouch Microcirculation

Background and purpose Propionyl-l-carnitine (pLc) exerts protective effects in different experimental models of ischemia–reperfusion (I/R). The aim of the present study was to assess the effects of intravenously and topically applied pLc on microvascular permeability increase induced by I/R in the hamster cheek pouch preparation. Methods The hamster cheek pouch microcirculation was visualized by fluorescence microscopy. Microvascular permeability, leukocyte adhesion to venular walls, perfused capillary length, and capillary red blood cell velocity (VRBC) were evaluated by computer-assisted methods. E-selectin expression was assessed by in vitro analysis. Lipid peroxidation and reactive oxygen species (ROS) formation were determined by thiobarbituric acid-reactive substances (TBARS) and 2′-7′-dichlorofluorescein (DCF), respectively. Results In control animals, I/R caused a significant increase in permeability and in the leukocyte adhesion in venules. Capillary perfusion and VRBC decreased. TBARS levels and DCF fluorescence significantly increased compared with baseline. Intravenously infused pLc dose-dependently prevented leakage and leukocyte adhesion, preserved capillary perfusion, and induced vasodilation at the end of reperfusion, while ROS concentration decreased. Inhibition of nitric oxide synthase prior to pLc caused vasoconstriction and partially blunted the pLc-induced protective effects; inhibition of the endothelium-derived hyperpolarizing factor (EDHF) abolished pLc effects. Topical application of pLc on cheek pouch membrane produced the same effects as observed with intravenous administration. pLc decreased the E-selectin expression. Conclusions pLc prevents microvascular changes induced by I/R injury. The reduction of permeability increase could be mainly due to EDHF release induce vasodilatation together with NO. The reduction of E-selectin expression prevents leukocyte adhesion and permeability increase

Long term remodeling of rat pial microcirculation after transient middle cerebral artery occlusion and reperfusion.

Objective: The aim of this study was to assess the in vivo structural and functional remodeling of pial arteriolar networks in the ischemic area of rats submitted to transient middle cerebral artery occlusion (MCAO) and different time intervals of reperfusion. Methods and results: Two closed cranial windows were implanted above the left and right parietal cortex to observe pial microcirculation by fluorescence microscopy. The geometric characteristics of pial arteriolar networks, permeability increase, leukocyte adhesion and capillary density were analyzed after 1 h or 1, 7, 14 or 28 days of reperfusion. MCAO and 1-hour reperfusion caused marked microvascular changes in pial networks. The necrotic core was devoid of vessels, while the penumbra area presented a few arterioles, capillaries and venules with severe neuronal damage. Penumbra microvascular permeability and leukocyte adhesion were pronounced. At 7 days of reperfusion, new pial arterioles were organized in anastomotic vessels, overlapping the ischemic core and in penetrating pial arterioles. Vascular remodeling caused different arteriolar rearrangement up to 28 days of reperfusion and animals gradually regained their motor and sensory functions. Conclusions: Transient MCAO-induced pial-network remodeling is characterized by arteriolar anastomotic arcades. Remodeling mechanisms appear to be accompanied by an increased expression of nitric oxide synthases

Hypotensive and bradycardic effects of mouth opening: evidence in the human

Objective Previous studies in normotensive anesthetized rats (Lapi Arch.Ital.Biol 151:11-23,2013) showed that peripheral stimulation of the trigeminal nerve induced by submaximal mouth opening (mandibular extension, ME) caused prolonged (at least 80min) bradycardia, hypotension and cerebral hemodynamic changes (pial arterioles showed a characteristic response pattern consisting in a significant constriction during ME followed by a dilatation for the entire remaining observation time). Design and method In this study we assessed the in vivo effects of ME on HR, MABP and pial microcirculation in hypertensive rats. Experiments were performed in male Wistar rats weighing 250-300g (n = 8). Hypertension was induced by intraperitoneal daily injection of dexamethasone (0.03mg/kg/day) for 10 days. ME was obtained by inserting an ad hoc developed retractor between the dental arches. HR and MABP were recorded by ECG and a catheter placed in the left femoral artery and measured by a computer-assisted system. Pial arterioles were observed through a closed cranial window implanted above the left parietal cortex and visualized by an in vivo fluorescence microscopy technique to assess vessel diameter changes before (baseline), during 10min ME and thereafter until 160min. Arteriolar diameters were measured with a computer-assisted method (MIP Image program, frame by frame). Results In sham-treated (no ME) hypertensive rats (n=3) HR, MABP and pial microcirculation did not change during whole observation period. Hypertensive rats subjected to ME (n=5) showed a significant decrease of HR and MABP. HR declined by 42bpm, (p<0.01) starting from 60 min after ME up to 160min, while MABP by 18mmHg (p<0.05) starting from 20min after ME up to 100min, compared with baseline. Pial arterioles exhibited a biphasic response: the arteriolar diameter decreased by 2.94&#956;m (p<0.05) during ME, afterwards it significantly increased by 3.46&#956;m (p<0.01) starting from 20min after ME; this vasodilatation lasted for the whole observation period. Conclusions Our results suggest that ME is able to exert profound and prolonged regulatory effects on systemic arterial blood pressure and pial arteriolar tone in hypertensive rats

Repeated mandibular extension in rat: A procedure to modulate the cerebral arteriolar tone

Previous data have shown both in the rat and in the human that a single mandibular extension lasting 10 min induces a significant important and prolonged reduction in blood pressure and heart rate, affecting also rat pial microcirculation by the release of endothelial factors. In the present work, we assessed whether repeated mandibular extension could further prolong these effects. We performed two mandibular extensions, the second mandibular extension being applied 10 min after the first one. The second mandibular extension produced a reduction in blood pressure and heart rate for at least 240 min. As in the case of a single mandibular extension, pial arterioles dilated persisting up to 140 min after the second extension. Spectral analysis on 30 min recordings under baseline conditions and after repetitive mandibular extensions showed that the pial arterioles dilation was associated with rhythmic diameter changes sustained by an increase in the frequency components related to endothelial, neurogenic, and myogenic activity while a single mandibular extension caused, conversely, an increase only in the endothelial activity. In conclusion, repetitive mandibular extension prolonged the effects of a single mandibular extension on blood pressure, heart rate and vasodilation and induced a modulation of different frequency components responsible of the pial arteriolar tone, in particular increasing the endothelial activity

Association of the body adiposity index (BAI) with metabolic risk factors in young and older overweight and obese women.

PURPOSE: Body adiposity index (BAI) is a novel index for the assessment of percentage fat mass (FM%). We tested the association between BAI and metabolic outcomes in overweight and obese women of different ages. METHODS: 260 young women (24.7 ± 5.3 years, 31.0 ± 5.0 kg/m(2)) and 328 older women (66.9 ± 4.6 years, 34.8 ± 4.7 kg/m(2)) were recruited. BAI was calculated using hip circumference and height. Bioimpedance analysis was used to measure FM%. Metabolic risk was assessed using a composite z score integrating standardised measurements of fasting glucose, total cholesterol, liver enzymes and triglycerides. RESULTS: The association between BAI and FM% was modest in both young (r = 0.56, p < 0.001) and older (r = 0.49, p < 0.001) groups. BAI was directly associated with metabolic risk in young women (r = 0.29, p < 0.001), whereas it showed a weak, inverse association in the older group (r = -0.14, p = 0.01). CONCLUSIONS: BAI validity needs to be re-assessed in older individuals for better definition of its predictive accuracy

Trigeminocardiac reflex by mandibular extension on rat pial microcirculation: Role of nitric oxide

In the present study we have extended our previous findings about the effects of 10 minutes of passive mandibular extension in anesthetized Wistar rats. By prolonging the observation time to 3 hours, we showed that 10 minutes mandibular extension caused a significant reduction of the mean arterial blood pressure and heart rate respect to baseline values, which persisted up to 160 minutes after mandibular extension. These effects were accompanied by a characteristic biphasic response of pial arterioles: during mandibular extension, pial arterioles constricted and after mandibular extension dilated for the whole observation period. Interestingly, the administration of the opioid receptor antagonist naloxone abolished the vasoconstriction observed during mandibular extension, while the administration of Nω-Nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, abolished the vasodilation observed after mandibular extension. Either drug did not affect the reduction of mean arterial blood pressure and heart rate induced by mandibular extension. By qRT-PCR, we also showed that neuronal nitric oxide synthase gene expression was significantly increased compared with baseline conditions during and after mandibular extension and endothelial nitric oxide synthase gene expression markedly increased at 2 hours after mandibular extension. Finally, western blotting detected a significant increase in neuronal and endothelial nitric oxide synthase protein expression. In conclusion mandibular extension caused complex effects on pial microcirculation involving opioid receptor activation and nitric oxide release by both neurons and endothelial vascular cells at different times