Studies in humans have found consumption of certain flavanoid-containing foods to be associated with improvement in endothelial function and with reduction in blood pressure. In particular, catechin, a natural phenol contained in cocoa and green tea, appears to play a crucial role in the prevention of cardiovascular diseases reducing the systemic arterial blood pressure. The aim of the present study was to in vivo investigate the effects of oral administration of catechin on pial arteriolar rhythmic diameter changes in SHR in comparison with normotensive rats. Catechin was orally administered to SHR for 2 months at the dosage of 30 mg/kg b.w. The pial microcirculation was visualized by fluorescence microscopy. The vessel diameter and rhythmic variations in diameter were evaluated with computer-assisted method. Moreover, the pial arterioles were classified by Strahler scheme in five orders. On 30 min of recordings was performed the Spectral analysis in order to evaluate different frequency components: the first two in the ranges 0.005-0.0095 Hz and 0.0095-0.021 Hz (endothelial activity), the second in the range 0.021-0.052 Hz (neurogenic activity), the third in the range 0.052-0.145 Hz (myogenic activity), finally, the fourth and the fifth in the ranges 0.145-0.6 Hz (respiratory activity) and 0.6-3.0 Hz (heart beat), respectively. The data indicate that SHR treated with catechin presented an increase in the amplitude frequency components related to endothelial activity that in untreated SHR were markedly reduced. In conclusion, the catechin treatment induces the recovery of endothelial activity in SHR where this frequency component was seriously compromised compared to normotensive rats.

The present study was aimed to assess the changes in skin microvascular blood flow (SBF) in newly diagnosed hyperglycemic obese subjects, administered with hypocaloric diet. Adult patients were recruited and divided in three groups: NW group (n=54), NG (n=54) and HG (n=54) groups were constituted by normal weight, normoglycemic and hyperglycemic obese subjects, respectively. SBF was measured by laser Doppler perfusion monitoring technique and oscillations in blood flow were analyzed by spectral methods under baseline conditions, at 3 and 6 months of dietary treatment. Under resting conditions, SBF was lower in HG group than in NG and NW ones. Moreover, all subjects showed blood flow oscillations with several frequency components. In particular, hyperglycemic obese patients revealed lower spectral density in myogenic-related component than normoglycemic obese and normal weight ones. Moreover, post-occlusive reactive hyperemia (PORH) was impaired in hyperglycemic obese compared to normoglycemic and normal weigh subjects. After hypocaloric diet, in hyperglycemic obese patients there was an improvement in SBF accompanied by recovery in myogenic-related oscillations and arteriolar responses during PORH. In conclusion, hyperglycemia markedly affected peripheral microvascular function; hypocaloric diet ameliorated tissue blood flow.

We previously reported that in normotensive humans submaximal mouth opening (mandibular extension) obtained by an ad hoc dilator (spring device), associated with partial masticatory movements and prolonged for 10 minutes is followed by a long-lasting reduction of blood pressure (BP) and heart rate (HR). Similar results were obtained by us in anesthetized rats. A recent independent study failed to confirm the results in the normotensive human. We reassessed, in 25 normotensive volunteers, the effects on BP and HR of mandibular extension obtained by the spring device associated with partial masticatory movements compared to a control procedure, consisting in keeping a tongue depressor between the incisor teeth. Both procedures were applied for 10 minutes and systolic BP (SBP), diastolic BP (DBP) and HR were measured every 10 minutes by an automatic recorder, for 30 minutes before and 120 minutes after the procedures in seated subjects watching nature documentary films on laptop screen.Baseline levels (mean of the last 3 measurements before procedure) did not significantly differ between the experimental and control sessions. Two way repeated measures ANOVA on absolute (recorded) values did not reveal a significant main effect of treatment for SBP, DBP and HR, but a significant main effect of time (P<0.001) for BP and HR. In addition, a significant interaction of time and treatment was found for SBP (P<0.001) and DBP (P=0.005), but not for HR. In addition, two way repeated measures ANOVA was done on changes from baseline obtaining a significant main effect of treatment (P<0.001) and time (P<0.001) and a significant interaction of time and treatment for SBP (P<0.001) and DBP (P<0.01). Post-hoc comparisons revealed significantly lower values for SBP and DBP in experimental compared to control values at almost all times and this decrement was by about 5 mmHg. Furthermore, for both absolute values and changes from baseline, the interaction effect was, for BP, of a qualitative type as indicated by an opposite effect in the time-course between control and experimental sessions. This study thus provides confirmatory evidence that submaximal mouth opening for a relatively brief time is followed by prolonged albeit small reductions of BP in normotensive human volunteers.

Microvascular dysfunctions are responsible for the vascular complications of diabetes, frequently observed in obese subjects. The present study was aimed to assess microvascular skin blood flow and related oscillations in hyperglycemic obese subjects.Twenty-eight normoglycemic controls and twenty-nine hyperglycemic subjects were recruited from our Outpatient Clinics. Anthropometric measurements were evaluated and bioimpedance analysis was performed. Moreover, glycemia, glycated hemoglobin (HbA1c) and basal insulin were measured. Finally, skin microvascular blood flow was determined by a laser Doppler flowmeter and oscillations in flow were analyzed by power spectrum methods.The mean skin blood flow and total power spectral density were lower in the hyperglycemic patients than in normoglycemic controls. Moreover, hyperglycemic subjects showed lower power spectral density of oscillations, related to myogenic activity, compared to the those observed in normoglycemic people.Therefore, hyperglycemia appears to be associated to impaired vascular smooth muscle cell function and alterations in microvascular blood flow regulation.

Hypertension causes several microvascular alterations, leading to inadequate blood supply to tissues and target-organ damage. The present study was aimed to investigate microvascular skin blood flow and related oscillations in hypertensive obese subjects. Twenty-eight controls and twenty-seven hypertensive patients were recruited from our Outpatient Clinics. Anthropometric measurements were evaluated and bioimpedance analysis was performed. Finally, skin microvascular blood flow was measured by a laser Doppler flowmeter and oscillations in blood flow were analyzed by power spectrum analysis. Newly diagnosed hypertensive subjects showed lower mean skin blood flow as well as lower related total power spectral density compared to the controls. In these patients, moreover, power spectral analysis of microvascular oscillations indicate a lower spectral density of the frequency component related to myogenic activity compared to controls. Therefore, hypertension appears to be associated to myogenic impairments, causing changes in microvascular modulation and tissue perfusion.

The aim of this study was to in vivo assess the effects of mandibular extension (ME) on rat pial rhythmic diameter changes. ME was induced by U-shaped device placed between the two dental arches. Pial microcirculation was visualized by fluorescence microscopy. The vessel diameter and rhythmic variations in diameter were evaluated with computer-assisted method. Pial arterioles were classified by Strahler method in five orders. Spectral analysis was performed on 30 min recordings under baseline conditions and 30 min tracings after 10 min ME. Under baseline conditions all pial arteriolar orders showed high and low frequency components; the low ones were in the ranges 0.0095-0.021 Hz (endothelial activity), 0.021-0.052 Hz (neurogenic activity), and 0.052-0.145 Hz (myogenic activity), 0.145-0.6 Hz (respiratory activity) and 0.6-2.0 Hz (heart beat). The smallest arterioles showed another low frequency component in the range 0.005-0.0095 Hz. After ME there was a significant increase in the lowest frequency components accompanied by a significant reduction in arteriolar blood pressure and heart rate. In conclusion, 10 min ME induces systemic responses and an increase in cerebral blood flow oscillations triggered by endothelial activity.

The present study was aimed to assess blood flow oscillatory patterns in rat pial microcirculation by laser speckle imaging. Laser speckle methods provides maps of cerebral blood flow with elevated temporal and spatial resolution. The study was carried out on male Wistar rats. An open cranial window was prepared on the parietal region. Pial arterioles were classified by Strahler method in five orders by fluorescence microscopy. The blood flow oscillatory patterns were determined by laser speckle imaging in single pial vessels. Power spectrum analysis was performed by wavelet methods under baseline conditions. Arterioles and venules were characterized by blood flow oscillations with frequency components in the ranges 0.005-0.0095 Hz, 0.0095-0.02 Hz, 0.02-0.15 Hz, 0.15-0.5 Hz. Arterioles showed oscillations with higher total power and higher spectral density in the range 0.02-0.15 when compared with venules. Laser speckle imaging allowed us to evaluate arteriolar and venular blood flow oscillations.

The trigemino-cardiac reflex is a brainstem reflex known to lead to a decrement in heart rate and blood pressure, whereas few data have been collected about its effects on the cerebral hemodynamic. In this study we assess the in vivo effects of trigeminal nerve peripheral stimulation by mandibular extension on pial microcirculation and systemic arterial blood pressure in rats. Experiments were performed in male Wistar rats subjected to mandibular extension obtained inserting an ad hoc developed retractor between the dental arches. Mean arterial blood pressure and heart rate were recorded and the pial arterioles were visualized by fluorescence microscopy to measure the vessel diameters before (15 minutes) during (5-15 minutes) and after (80 minutes) mandibular extension. While in control rats (sham-operated rats) and in rats subjected to the dissection of the trigeminal peripheral branches mean arterial blood pressure, heart rate and pial microcirculation did not change during the whole observation period (110 minutes), in rats submitted to mandibular extension, mean arterial blood pressure, heart rate and arteriolar diameter significantly decreased during stimulation. Afterward mean arterial blood pressure remained reduced as well as heart rate, while arteriolar diameter significantly increased evidencing a vasodilatation persisting for the whole remaining observation time. Therefore, trigeminal nerve proprioceptive stimulation appears to trigger specific mechanisms regulating systemic arterial blood pressure and pial microcirculation.

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.

The aim of the present study was to assess quercetin's mechanism of action in rat pial microvessels during transient bilateral common carotid artery occlusion (BCCAO) and reperfusion. Rat pial microcirculation was visualized using fluorescence microscopy through a closed cranial window. Pial arterioles were classified in five orders of branchings. In ischemic rats, 30 min BCCAO and 60 min reperfusion caused arteriolar diameter decrease, microvascular leakage, leukocyte adhesion in venules, and reduction of capillary perfusion. Quercetin highest dose determined dilation in all arteriolar orders, by 40 ± 4% of baseline in order 2 vessels, and prevented microvascular permeability [0.15 ± 0.02 normalized gray levels (NGL)], leukocyte adhesion, and capillary failure. Protein kinase C (PKC) inhibition exerted by chelerythrine prior to quercetin attenuated quercetin-induced effects: order 2 arterioles dilated by 19.0 ± 2.4% baseline, while there was an increase in permeability (0.40 ± 0.05 NGL) and leukocyte adhesion with a marked decrease in capillary perfusion. Tyrosine kinase (TK) inhibition by tyrphostin 47 prior to quercetin lessened smaller pial arterioles responses, dilating by 20.7 ± 2.5% of baseline, while leakage increased (0.39 ± 0.04 NGL) sustained by slight leukocyte adhesion and ameliorated capillary perfusion. Inhibition of endothelium nitric oxide synthase (eNOS) by N(G)-nitro-L-arginine-methyl ester (L-NAME) prior to PKC or TK reduced the quercetin's effects on pial arteriolar diameter and leakage. eNOS inhibition by L-NAME reduced quercetin effects on pial arteriolar diameter and leakage. Finally, combined inhibition of PKC and TK prior to quercetin abolished quercetin-induced effects, decreasing eNOS expression, while blocking ATP-sensitive potassium (K(ATP)) channels by glibenclamide suppressed arteriolar dilation. In conclusion, the protective effects of quercetin could be due to different mechanisms resulting in NO release throughout PKC and TK intracellular signaling pathway activation.

Various procedures involving stimulations of facial regions are known to induce so-called trigemino-cardiac reflexes that entail a decrease of heart rate and blood pressure. We here report the effects of a specific stimulation that consists in a submaximal passive mandibular extension obtained by means of a dilatator applied for 10 minutes between the upper and lower incisor teeth, associated with partial active masticatory movements. Blood pressure and heart rate were determined in 18 young normal volunteers by a Omron M4 recorder, before (20 minutes), during (10 minutes) and after mandibular extension (80 minutes) and under control conditions (same overall duration without stimulation). While control values remained stable, mandibular extension was followed by a progressive decline of both blood pressure (up to about 12/11 mmHg) and heart rate (up to about 13 bpm), statistically confirmed by ANOVA both on absolute values and on changes from basal values. The decline of systolic blood pressure and heart rate significantly correlated with basal values. The present findings indicate that submaximal opening of the mouth, associated to partial masticatory movements, induces a prolonged reduction of blood pressure and heart rate in normotensive volunteers.

Objective: The aim of the study was to assess the rat pial microvessel alterations due to transient bilateral common carotid artery occlusion (BCCAO) and to investigate the mechanism of 10% hypertonic glycerol neuroprotection. Our suggestion was that 10% glycerol solution infusion could dilate pial arterioles through nitric oxide release and/or stimulation of ATP-sensitive potassium (KATP) channels. Therefore, we studied the effects of hypertonic glycerol after inhibition of nitric oxide synthase, with NG-nitro-L-arginine-methyl ester or NG-nitro-L-arginine, or KATP channels with glibenclamide. Methods: Pial microcirculation of male Wistar rats was visualized by a fluorescent microscopy technique through an open cranial window, using fluorescein isothiocyanate bound to dextran (molecular weight 70 kDa). BCCAO was induced for 30 min and reperfusion lasted 60 min. The arterioles were classified according to the Strahler ordering scheme. Permeability increase was quantified by normalized grey levels (NGL). Leucocytes were stained with rhodamine 6G. Perfused capillary length and capillary red blood cell (RBC) velocity were measured by computer-assisted methods. Results: The arterioles were assigned 5 orders of branchings, from order 1 (diameter 16.0 ± 2.5 µm) to order 5 (62.0 ± 5.0 µm). BCCAO caused inhomogenous changes in diameter of arterioles and leakage of fluorescent dextran, that was further enhanced by reperfusion (0.45 ± 0.05 NGL, p < 0.01). Adhesion of leukocytes to venules was marked and capillary perfusion was reduced by 39.2 ± 6.0% of baseline as well as capillary RBC velocity. 10% glycerol solution caused an increase in diameter of all arterioles within 25 ± 2 min of administration (by 20 ± 5% in order 4, 25 ± 4% in order 3 and 18 ± 3% in order 2; p < 0.01). Leakage (0.19 ± 0.03 NGL, p < 0.01), leukocyte adhesion (2.0 ± 1.0/100 µm of venular length, p < 0.01) and capillary occlusion (reduction by 13.0 ± 5.5% of baseline) were prevented compared with controls. Capillary RBC velocity increased compared with controls. NG-nitro-L-arginine-methyl ester or NG-nitro-L-arginine infused prior to glycerol caused vasoconstriction and reduced the protective effects of hypertonic glycerol on permeability increase. The number of adherent leukocytes and perfused capillary length decreased, while capillary RBC velocity was higher than baseline. Glibenclamide prior to 10% glycerol solution blunted glycerol-induced vasodilatation, but did not affect protection by hypertonic glycerol on blood-brain barrier disruption, leukocyte adhesion and capillary perfusion, preserving high capillary RBC velocity. Papaverine (20 mg/kg body weight) induced an increase in arteriolar diameter, enhancing interstitial edema; adhesion of leukocytes was marked as well as capillary occlusion, while capillary RBC velocity increased. Conclusions: 10% glycerol solution was able to prevent microvascular alterations due to BCCAO protecting cerebral tissue. The effects appear to be due to hyperosmolality causing stimulation of KATP channels, increase in vessel wall shear stress and release of nitric oxide.