Ana Cristina Martínez

Neuronal and smooth muscle receptors involved in the PACAP- and VIP-induced relaxations of the pig urinary bladder neck

As pituitary adenylate cyclase‐activating polypeptide 38 (PACAP 38)‐ and vasoactive intestinal peptide (VIP) are widely distributed in the urinary tract, the current study investigated the receptors and mechanisms involved in relaxations induced by these peptides in the pig bladder neck.

Blunted acetylcholine relaxation and nitric oxide release in arteries from renal hypertensive rats.

Objective Investigation of the effect of hypertension on endothelium-dependent relaxation and release of nitric oxide (NO) in normotensive and renal hypertensive rats. Design and methods Sprague-Dawley rats were randomly allocated into two groups: uninephrectomized controls and one-kidney one-clip (Goldblatt hypertension) hypertensive rats, a non-renin dependent model of hypertension. After 10 weeks and in the presence of the cyclooxygenase inhibitor indomethacin, simultaneous measurements of the NO concentration, measured with a NO-specific microelectrode and endothelium-dependent relaxation were performed in isolated rat superior mesenteric arteries. Results Addition of the NO scavenger, oxyhaemoglobin, showed that basal NO concentration was unaltered in arterial segments from hypertensive rats. In norepinephrine-contracted arteries, acetylcholine increased the NO concentration and caused relaxations, and both parameters were significantly reduced in renal hypertensive arteries. Relaxations induced by the NO donor, S-nitroso-N-acetylpenicillamine were reduced. The superoxide scavenger, superoxide dismutase, and the NO synthase substrate, l-arginine, did not change the increase in NO concentration or acetylcholine relaxation in arteries from normotensive or renal hypertensive animals. In contrast, the NO synthase inhibitor, asymmetric dimethyl l-arginine, reduced the NO concentration and acetylcholine relaxation, while these responses were abolished in the presence of oxyhaemoglobin. Conclusions This study provides direct evidence that reduced endothelium-dependent relaxations in the superior mesenteric artery from renal hypertensive rats is due, at least in part, to diminished NO release. The reduced NO release and relaxation persist in the presence of excess of substrate for NO synthase.

Mechanisms involved in testosterone-induced vasodilatation in pig prostatic small arteries

AIMS Testosterone is beneficial to the cardiovascular system due to its direct coronary vasodilatory action and its circulatory deficiency is associated with coronary artery disease (CAD), which has been proposed as an extrinsic risk factor for benign prostatic hyperplasia (BPH). Therefore, the current study investigated the mechanisms involved in the testosterone-induced vasodilatation in pig prostatic small arteries. MAIN METHODS The testosterone vasoactive effects were assessed in small arterial rings mounted in microvascular myographs for isometric force recordings. KEY FINDINGS Testosterone and the non-aromatizable metabolite 4, 5alpha-dihydrotestosterone (DHT) evoked a similar concentration-dependent relaxation on noradrenaline (NA)-precontracted rings. Similar responses were obtained in preparations contracted with 60 mM K(+)-enriched physiological saline solution. Endothelium mechanical removal or pre-treatment with blockers of nitric oxide (NO) synthase, guanylate cyclase, aromatase activity, intracellular androgenic receptor (AR), 5alpha-reductase, prostanoid synthesis and K(+) channels, failed to modify the responses to testosterone. In Ca(2+)-free 124 mM KPSS, testosterone markedly inhibited in a concentration-dependent manner the contraction curve t degrees CaCl(2). In arteries pretreated with an L-type voltage-activated Ca(2+) channels (VOCCs) inhibitor, nifedipine, testosterone still relaxed noradrenaline-precontracted arteries. SIGNIFICANCE These data suggest that testosterone induces a direct vasodilatory action in pig prostatic small arteries independent of either endothelium, NO, prostanoids, aromatase or 5alpha-reductase activities, AR or K(+) channels. Such an effect is suggested to be produced via blockade of extracellular Ca(2+) entry through L-type VOCCs and non-L-type Ca(2+) channels. Testosterone-induced vasodilatation could be useful to prevent prostatic ischemia.

5-hydroxytryptamine induced relaxation in the pig urinary bladder neck

Background and purpose:  5‐Hydroxytryptamine (5‐HT) is one of the inhibitory mediators in the urinary bladder outlet region. Here we investigated mechanisms involved in 5‐HT‐induced relaxations of the pig bladder neck.

Pre-junctional α2-adrenoceptors modulation of the nitrergic transmission in the pig urinary bladder neck†

AIMS To investigate the nitric oxide (NO)-mediated nerve relaxation and its possible modulation by pre-junctional alpha2-adrenoceptors in the pig urinary bladder neck. METHODS Urothelium-denuded bladder neck strips were dissected, and mounted in isolated organ baths containing a physiological saline solution (PSS) at 37 degrees C and continuously gassed with 5% CO2 and 95% O2, for isometric force recording. The relaxations to transmural nerve stimulation (electrical field stimulation [EFS]) or exogenously applied NO were carried out on strips pre-contracted with 1 microM phenylephrine (PhE) and treated with guanethidine (10 microM) and atropine (0.1 microM), to block noradrenergic neurotransmission and muscarinic receptors, respectively. RESULTS EFS (0.2-1 Hz, 1 msec duration, 20 sec trains, current output adjusted to 75 mA) evoked frequency-dependent relaxations which were abolished by the neuronal voltage-activated Na+ channel blocker tetrodotoxin (TTX, 1 microM). These responses were potently reduced by the nitric oxide synthase (NOS) inhibitor NG-nitro-L-arginine (L-NOARG, 30 microM) and further reversed by the NO synthesis substrate L-arginine (L-ARG, 3 mM). The alpha2-adrenoceptor agonist BHT-920 (2 microM) reduced the electrically evoked relaxations, its effectiveness being higher on the responses induced by low frequency stimulation. BHT-920-elicited reductions were fully reversed by the alpha2-adrenoceptor antagonist rauwolscine (RAW, 1 microM). Exogenous NO (1 microM-1 mM) induced concentration-dependent relaxations which were not modified by BHT-920, thus eliminating a possible post-junctional modulation. CONCLUSIONS These results indicate that NO is involved in the non-adrenergic non-cholinergic (NANC) inhibitory neurotransmission in the pig urinary bladder neck, the release of NO from intramural nerves being modulated by pre-junctional alpha2-adrenoceptor stimulation.

Altered arachidonic acid metabolism via COX‐1 and COX‐2 contributes to the endothelial dysfunction of penile arteries from obese Zucker rats

Background and purpose:  The aim of the current study was to investigate the role of arachidonic acid (AA) metabolism via cyclooxygenase (COX) in the endothelial dysfunction of penile arteries from pre‐diabetic, obese Zucker rats (OZR).

Mechanisms of the relaxant effect of vardenafil in rat penile arteries

The aim of the present study was to investigate the mechanisms underlying the vasorelaxation induced by the selective phosphodiesterase 5 (PDE5) inhibitor vardenafil in rat penile small arteries. Segments of the rat dorsal penile artery were mounted in microvascular myographs for isometric tension recording. Concentration-response curves for vardenafil (1 nM-3 microM) and other PDE inhibitors (sildenafil, rolipram and milrinone) were constructed by adding cummulative concentrations of the drugs to arteries precontracted with phenylephrine. The effect of mechanical endothelial cell removal and of selective blockers of the nitric oxide (NO)/cGMP pathway and K+ channels were evaluated on the vardenafil relaxant responses. Vardenafil was the most potent of the four PDE inhibitors tested that maximally relaxed penile arteries, pD2 and maximum relaxation being 6.96+/-0.08 and 97+/-1% (n=48), respectively. Blockade of guanylate cyclase with ODQ (5 microM), mechanical removal of the endothelium or inhibition of NO synthase with l-NOARG (100 microM) markedly reduced vardenafil-induced relaxations, without altering maximum response. Inhibitors of both the cGMP-dependent (PKG) and the cAMP-dependent (PKA) protein kinases, Rp-8-Br-PET-cGMPS (5 microM) and Rp-8-CPT-cAMPS (50 microM), respectively, both reduced vardenafil relaxant responses and the later abolished that of rolipram. Vardenafil-elicited relaxation was reduced by the selective inhibitor of the large-conductance Ca2+-activated K+ channels (BK(Ca)), iberiotoxin (30 nM) and also by the ATP-sensitive K+ channel (K(ATP)) inhibitor, glibenclamide (1 microM). Vardenafil induces a potent vasodilatation in rat penile arteries that is partially dependent on the endothelium and the NO/cGMP pathway and involves activation of both BK(Ca) and K(ATP) channels.

Heterogeneity of neuronal and smooth muscle receptors involved in the VIP- and PACAP-induced relaxations of the pig intravesical ureter

The mechanisms and receptors involved in the vasoactive intestinal peptide (VIP)‐ and pituitary adenylate cyclase‐activating polypeptide (PACAP)‐induced relaxations of the pig intravesical ureter were investigated. VIP, PACAP 38 and PACAP 27 concentration‐dependently relaxed U46619‐contracted ureteral strips with a similar potency. [Ala11,22,28]‐VIP, a VPAC1 agonist, showed inconsistent relaxations. The neuronal voltage‐gated Ca2+ channel inhibitor, ω‐conotoxin GVIA (ω‐CgTX, 1 μM), reduced the VIP relaxations. Urothelium removal or blockade of capsaicin‐sensitive primary afferents, nitric oxide (NO) synthase and guanylate cyclase with capsaicin (10 μM), NG‐nitro‐L‐arginine (L‐NOARG, 100 μM) and 1H‐[1,2,4]‐oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ, 5 μM), respectively, did not change the VIP relaxations. However, the PACAP 38 relaxations were reduced by ω‐CgTX, capsaicin, L‐NOARG and ODQ. The VIP and VIP/PACAP receptor antagonists, [Lys1, Pro2,5, Arg3,4, Tyr6]‐VIP (1 μM) and PACAP (6–38) (0.4 μM), inhibited VIP and VIP and PACAP 38, respectively, relaxations. The nonselective and large‐conductance Ca2‐activated K+ channel blockers, tetraethylammonium (3 mM) and charybdotoxin (0.1 μM), respectively, and neuropeptide Y (0.1 μM) did not modify the VIP relaxations. The small‐conductance Ca2‐activated K+ channel blocker apamin (1 μM) did not change the PACAP 27 relaxations. The cAMP‐dependent protein kinase A (PKA) blocker, 8‐(4‐chlorophenylthio)adenosine‐3′,5′‐cyclic monophosphorothioate (Rp‐8‐CPT‐cAMPS, 100 μM), reduced VIP relaxations. The phosphodiesterase 4 inhibitor rolipram and the adenylate cyclase activator forskolin relaxed ureteral preparations. The rolipram relaxations were reduced by Rp‐8‐CPT‐cAMPS. Forskolin (30 nM) evoked a potentiation of VIP relaxations. These results suggest that VIP and PACAP relax the pig ureter through smooth muscle receptors, probably of the VPAC2 subtype, linked to a cAMP‐PKA pathway. Neuronal VPAC receptors localized at motor nerves and PAC1 receptors placed at sensory nerves and coupled to NO release, seem also to be involved in the VIP and PACAP 38 relaxations.

Characterization of the 5‐hydroxytryptamine receptors mediating contraction in the pig isolated intravesical ureter

This study was designed to investigate the effect of 5‐hydroxytryptamine (5‐HT) and to characterize the 5‐HT receptors involved in 5‐HT responses in the pig intravesical ureter. 5‐HT (0.01–10 μM) concentration‐dependently increased the tone of intravesical ureteral strips, whereas the increases in phasic contractions were concentration‐independent. The 5‐HT2 receptor agonist α‐methyl 5‐HT, mimicked the effect on tone whereas weak or no response was obtained with 5‐CT, 8‐OH‐DPAT, m‐chlorophenylbiguanide and RS 67333, 5‐HT1, 5‐HT1A, 5‐HT3 and 5‐HT4 receptor agonists, respectively. 5‐HT did not induce relaxation of U46619‐contracted ureteral preparations. Pargyline (100 μM), a monoaminooxidase A/B activity inhibitor, produced leftward displacements of the concentration‐response curves for 5‐HT. 5‐HT‐induced tone was reduced by the 5‐HT2 and 5‐HT2A receptor antagonists ritanserine (0.1 μM) and spiperone (0.2 μM), respectively. However, 5‐HT contraction was not antagonized by cyanopindolol (2 μM), SDZ–SER 082 (1 μM), Y‐25130 (1 μM) and GR 113808 (0.1 μM), which are respectively, 5‐HT1A/1B, 5‐HT2B/2C, 5‐HT3, and 5‐HT4 selective receptor antagonists. Removal of the urothelium did not modify 5‐HT‐induced contractions. Blockade of neuronal voltage‐activated sodium channels, α‐adrenergic receptors and adrenergic neurotransmission with tetrodotoxin (1 μM), phentolamine (0.3 μM) and guanethidine (10 μM), respectively, reduced the contractions to 5‐HT. However, physostigmine (1 μM), atropine (0.1 μM) and suramin (30 μM), inhibitors of cholinesterase activity, muscarinic‐ and purinergic P2‐receptors, respectively, failed to modify the contractions to 5‐HT. These results suggest that 5‐HT increases the tone of the pig intravesical ureter through 5‐HT2A receptors located at the smooth muscle. Part of the 5‐HT contraction is indirectly mediated via noradrenaline release from sympathetic nerves.

Endothelium‐Independent Relaxation Induced By Histamine In Human Dorsal Penile Artery

1. In vitro preparations of human dorsal penile arteries were used to evaluate the effect of histamine and to characterize the histamine receptors involved in the response.