Róbert Pórszász

Tissue-Specific Regulation of Microvascular Diameter: Opposite Functional Roles of Neuronal and Smooth Muscle Located Vanilloid Receptor-1

The transient receptor potential type V1 channel (vanilloid receptor 1, TRPV1) is a Ca2+-permeable nonspecific cation channel activated by various painful stimuli including ischemia. We hypothesized that TRPV1 is expressed in the arterioles and is involved in the regulation of microvascular tone. We found that TRPV1 stimulation by capsaicin (intra-arterial administration) of the isolated, perfused right hind limb of the rat increased vascular resistance (by 98 ± 21 mm Hg at 10 μg) in association with decreased skeletal muscle perfusion and elevation of skin perfusion (detected by dual-channel laser Doppler flowmetry). Denervation of the hind limb did not affect capsaicin-evoked changes in vascular resistance and tissue perfusion in the hind limb but reduced the elevation of perfusion in the skin. In isolated, pressurized skeletal (musculus gracilis) muscle arterioles (diameter, 147 ± 35 μm), capsaicin had biphasic effects: at lower concentrations, capsaicin (up to 10 nM) evoked dilations (maximum, 32 ± 13%), whereas higher concentrations (0.1-1 μM) elicited substantial constrictions (maximum, 66 ± 7%). Endothelium removal or inhibition of nitric-oxide synthase abolished capsaicin-induced dilations but did not affect arteriolar constriction. Expression of TRPV1 was detected by reverse transcriptase-polymerase chain reaction in the aorta and in cultured rat aortic vascular smooth muscle cells (A7r5). Immunohistochemistry revealed expression primarily in the smooth muscle layers of the gracilis arteriole. These data demonstrate the functional expression of TRPV1 in vascular smooth muscle cells mediating vasoconstriction of the resistance arteries. Because of the dual effects of TRPV1 stimulation on the arteriolar diameter (dilation in skin, constriction in skeletal muscle), we propose that TRPV1 ligands represent drug candidates for tissue-specific modulation of blood distribution.

Vanilloid receptor-1 (TRPV1) expression and function in the vasculature of the rat.

Transient receptor potential (TRP) cation channels are emerging in vascular biology. In particular, the expression of the capsaicin receptor (TRPV1) was reported in vascular smooth muscle cells. This study characterized the arteriolar TRPV1 function and expression in the rat. TRPV1 mRNA was expressed in various vascular beds. Six commercially available antibodies were tested for TRPV1 specificity. Two of them were specific (immunostaining was abolished by blocking peptides) for neuronal TRPV1 and one recognized vascular TRPV1. TRPV1 was expressed in blood vessels in the skeletal muscle, mesenteric and skin tissues, as well as in the aorta and carotid arteries. TRPV1 expression was found to be regulated at the level of individual blood vessels, where some vessels expressed, while others did not express TRPV1 in the same tissue sections. Capsaicin (a TRPV1 agonist) evoked constrictions in skeletal muscle arteries and in the carotid artery, but had no effect on the femoral and mesenteric arteries or the aorta. In blood vessels, TRPV1 expression was detected in most of the large arteries, but there were striking differences at level of the small arteries. TRPV1 activity was suppressed in some isolated arteries. This tightly regulated expression and function suggests a physiological role for vascular TRPV1.

Hepatic insulin sensitizing substance: a novel ‘sensocrine’ mechanism to increase insulin sensitivity in anaesthetized rats

We recently described the sensory nitrergic nature of the hepatic insulin sensitizing substance (HISS) mechanism linked to postprandial activation of anterior hepatic plexus fibres in rabbits. This study is designed to assess the involvement of the sensory pathways in this mechanism. Selective sensory denervation of the anterior hepatic plexus (AHP) was achieved by a 3‐day perineurial treatment with 2% capsaicin solution in Wistar rats (230–250 g). After 1 week, hyperinsulinaemic (100 μU kg−1) euglycaemic (5.5 mmol kg−1) glucose clamp studies were performed to estimate insulin sensitivity. The rats with regional AHP sensory denervation exhibited a significantly decreased insulin sensitivity, that is, 9.1±1.0 mg kg−1 min−1 glucose reinstalled euglycaemia vs 13.3±1.9 mg kg−1 min−1 glucose (P<0.01) in control rats. Acute partial hepatic denervation by AHP cut was without effect on insulin sensitivity, whereas chronic hepatic denervation induced insulin resistance was similar to that achieved by regional AHP capsaicin treatment. Intraportal administration of L‐NAME (10 mg kg−1) decreased, whereas capsaicin (0.3 mg kg−1 min−1) increased insulin sensitivity. Neither atropine (1 mg kg−1) nor acetylcholine (1–10 μg mg min−1) produced any significant effect. In animals with preceding regional capsaicin desensitization, none of the pharmacological manoeuvres modified the resulting insulin‐resistant state. Cysteamine (200 mg kg−1 s.c.) is known to cause functional somatostatin depletion‐induced insulin resistance similar to that produced by either chronic partial hepatic denervation or perineurial AHP capsaicin desensitization. Intraportal capsaicin (0.3 mg kg−1 min−1) was unable to modify insulin resistance achieved by cysteamine. We conclude that capsaicin‐sensitive sensory fibres play a crucial role in neurogenic insulin sensitization known as the HISS mechanism without involvement of anatomical reflex‐mediated circuits. The results also suggest that HISS is identical to somatostatin of AHP sensory neural origin.

The sensory nitrergic nature of the hepatic insulin sensitizing substance mechanism in conscious rabbits

Functional deterioration of sensory fibres in the anterior hepatic plexus or intraportal administration of 7-nitro indazole (1 mg/kg), a selective inhibitor of neural nitric oxide (NO) synthase, caused insulin resistance as determined by hyperinsulinaemic (100 micro U/ml) euglycaemic (5.5 mmol/l) glucose clamping in chronically instrumented conscious rabbits. Intraportal nitroglycerin restored insulin sensitivity in either case. We conclude that NO of sensory neural origin plays a major role in endogenous neurogenic insulin sensitizing mechanisms.

Structure‐activity relationships of vanilloid receptor agonists for arteriolar TRPV1

BACKGROUND AND PURPOSE The transient receptor potential vanilloid 1 (TRPV1) plays a role in the activation of sensory neurons by various painful stimuli and is a therapeutic target. However, functional TRPV1 that affect microvascular diameter are also expressed in peripheral arteries and we attempted to characterize this receptor.

Capsaicin-induced nonneural vasoconstriction in canine mesenteric arteries

Prolonged cold storage (4 degrees C) of canine mesenteric arteries was used to reveal the role of nonneural mechanisms in capsaicin-induced vascular contraction. The EC(50) values of capsaicin were 3.0 microM, 670 and 104 nM in preparations made fresh, after a 1- or 2-week period of cold storage, respectively, indicating an enhanced contractile responsiveness of the denervated tissue to capsaicin. A similar exaggerated contractile response was seen with phenylephrine exclusively after a 1-week cold storage. For fresh, 1- and 2-week cold-stored arteries, the EC(50) of phenylephrine were 248, 38 and 30 nM, respectively. The maximum contraction produced by tyramine was decreased with time. The results suggest that capsaicin may attain vasoconstriction independent of neural elements.

Different desensitization patterns for sensory and vascular TRPV1 populations in the rat: Expression, localization and functional consequences

Background and purpose TRPV1 is expressed in sensory neurons and vascular smooth muscle cells, contributing to both pain perception and tissue blood distribution. Local desensitization of TRPV1 in sensory neurons by prolonged, high dose stimulation is re-engaged in clinical practice to achieve analgesia, but the effects of such treatments on the vascular TRPV1 are not known. Experimental approach Newborn rats were injected with capsaicin for five days. Sensory activation was measured by eye wiping tests and plasma extravasation. Isolated, pressurized skeletal muscle arterioles were used to characterize TRPV1 mediated vascular responses, while expression of TRPV1 was detected by immunohistochemistry. Key results Capsaicin evoked sensory responses, such as eye wiping (3.6±2.5 versus 15.5±1.4 wipes, p<0.01) or plasma extravasation (evans blue accumulation 10±3 versus 33±7 µg/g, p<0.05) were reduced in desensitized rats. In accordance, the number of TRPV1 positive sensory neurons in the dorsal root ganglia was also decreased. However, TRPV1 expression in smooth muscle cells was not affected by the treatment. There were no differences in the diameter (192±27 versus 194±8 µm), endothelium mediated dilations (evoked by acetylcholine), norepinephrine mediated constrictions, myogenic response and in the capsaicin evoked constrictions of arterioles isolated from skeletal muscle. Conclusion and implications Systemic capsaicin treatment of juvenile rats evokes anatomical and functional disappearance of the TRPV1-expressing neuronal cells but does not affect the TRPV1-expressing cells of the arterioles, implicating different effects of TRPV1 stimulation on the viability of these cell types.

Impairment by lovastatin of neural relaxation of the rabbit sphincter of Oddi

We sought whether inhibition of cholesterol biosynthesis by lovastatin influenced the nitrergic relaxation response of the sphincter of Oddi. Rabbit sphincters of Oddi rings were tested for changes in isometric tension in response to field stimulation in the presence of 4 microM guanethidine and 1 microM atropine. Tissue samples were then analyzed for cAMP and cGMP content by radioimmunoassay for nitric oxide concentration by electron spin resonance and for vasoactive intestinal peptide and calcitonin gene-related peptide (CGRP) release by radioimmunoassay. Membrane G(salpha) protein was determined by Western blot analysis. Field stimulation relaxed the preparations with an increase in nitric oxide, cAMP and cGMP concentrations at increased calcitonin gene-related peptide and vasoactive intestinal polypeptide (VIP) release. Preparations from rabbits pre-treated with lovastatin (5 mg/kg/day intragastrically, over 5 days) contracted under the same conditions with an attenuated cGMP-increase at preserved increase in NO content and neuropeptide release. The relaxation was recaptured combining lovastatin with farnesol (1 mg/kg intravenously, twice a day for 5 days). The field stimulation-induced increase in cyclic nucleotides was also restored. Lovastatin decreased membrane G(salpha) protein content, which was re-normalized by farnesol. Farnesol treatment reinstates neurogenic relaxation of the sphincter of Oddi deteriorated by lovastatin possibly by normalizing G-protein coupling.

Analgesic and anti-inflammatory effectiveness of sitagliptin and vildagliptin in mice.

To validate the potential anti-inflammatory and analgesic role of sita- and vildagliptin, five different experimental models were used in mice: i) mustard oil-induced ear edema, ii) neutrophil accumulation, iii) mechanical and iv) thermal touch sensitivity in complete Freunds adjuvant-induced arthritis and v) capsaicin-induced plasma extravasation in the urinary bladder. For the complete examination period in i) the dose of 10mg sitagliptin as well as 1-10mg vildagliptin was found to significantly decrease ear edema as compared to positive control (p<0.05, n=8/group). All doses of sitagliptin provided an anti-inflammatory effect p<0.005 (n=10/group) in test ii) and an analgesic effect in iii) except 3mg. Vildagliptin was similarly effective in test ii) (p<0.005, n=10/group) as sitagliptin, but it failed to affect mechanical touch sensitivity. Unlike mechanical touch sensitivity, both gliptins could beneficially act on the thermal threshold (p<0.05, n=10/group). And only in tests v) could both gliptins reverse inflammation. Further studies are needed to support the suggestion that the utilization of these beneficial effects of gliptins may be considered in the treatment of Type 2 diabetic patients.

Daily exposure to inhomogeneous static magnetic field significantly reduces blood glucose level in diabetic mice

Purpose: The present study was designed to reveal, whether the 30 min daily full body exposure of mice to an inhomogeneous static magnetic field (SMF) has a statistically significant effect on diabetic neuropathy. Materials and methods: Three different doses (100, 150, or 200 mg/kg) of streptozotocin (STZ) were administered intraperitoneally in groups of mice to induce diabetes. Body weight, blood glucose level and the nociceptive temperature threshold of mice were monitored for a period of 12 weeks. The group treated with 200 mg/kg i.p. STZ produced manifest diabetic neuropathy. Results were compared to non-treated (no SMF, no STZ) and SMF exposure without STZ-treatment group (SMF, no STZ) group. The inhomogeneous SMF was in the range of 2.8–476.7 mT peak-to-peak magnetic flux density. Results: Whereas SMF exposure did not seem to affect body weight and nociceptive temperature threshold, it statistically significantly (p < 0.001) reduced blood glucose level in the 200 mg/kg STZ (n = 6) group. Conclusions: Daily SMF exposure repeated for several weeks is protective against the development of high blood glucose level in diabetic mice.