Albino García-Sacristán

Penile arteries and erection.

Alterations in the flow of blood to and from the penis are thought to be the most frequent causes of male erectile dysfunction and, therefore, the present review focuses on the penile vasculature. In the flaccid state, tonic noradrenaline release from the sympathetic nerves contracts penile arterial and corporal smooth muscle through activation of postjunctional α1-adrenoceptors, both by increasing intracellular calcium and by enhancing the sensitivity of the contractile apparatus for calcium. In addition, noradrenaline inhibits vasodilatatory neurotransmitter release by prejunctional α2-adrenoceptors. The exact role of the sympathetic neurotransmitters, neuropeptide Y and adenosine 5′-triphosphate, in erection is largely unknown. Penile vasodilatation during erection is mediated by nitric oxide (NO) through activation of guanylyl cyclase in the smooth muscle layer, followed by increases in cyclic guanosine monophosphate lowering of intracellular calcium and desensitisation of the contractile apparatus for calcium. Acetylcholine, vasoactive intestinal peptide as well as peptides in sensory nerves probably also play a role in penile vasodilation. Increased flow through the penile arteries stimulates the endothelium leading to release of NO, prostanoids and a non-NO non-prostanoid factor, and as such enhances the vasodilatation, while the role of endothelium-derived contractile factors in penile vasoconstriction is not clear. Erectile dysfunction shares arterial risk factors with ischaemic heart disease, and diabetes, age, and hypercholesterolaemia are associated with impairment of both neurogenic and endothelium-dependent vasodilator mechanisms in corpus cavernosum. Only few studies have investigated the impact of these risk factors on the penile vasculature, although recent evidence suggests that arterial insufficiency precedes changes in corpus cavernosum leading to erectile dysfunction.

Noradrenaline modulates smooth muscle activity of the isolated intravesical ureter of the pig through different types of adrenoceptors

1 We have studied the effects of α‐ and β‐adrenoceptor agonists and antagonists on both phasic peristaltic activity and basal tone of the isolated intravesical ureter of the pig by means of isometric techniques in vitro. 2 Spontaneous phasic activity was exhibited by 21% of pig intravesical ureter preparations manifested as rhythmic contractions with average frequency and amplitude of 2.54 ± 0.18 min−1 and 1.48 ± 0.16 g (n = 31), respectively. 3 Adrenaline, noradrenaline and phenylephrine induced concentration‐dependent increases in both phasic activity and basal tone of ureteral preparations, all three agonists being more potent in modifying ureteral phasic activity than baseline tone. B‐HT 920, B‐HT 933 and clonidine had no significant effect. 4 Phentolamine (10−9 − 10−7 m) and prazosin (3 × 10−11 − 3 × 10−8 m) significantly inhibited increases in both frequency of phasic activity and baseline tone induced by a submaximal dose of noradrenaline. Rauwolscine (10−9 − 10−7 m) affected only the tone evoked by noradrenaline and higher concentrations of this antagonist were needed to block phasic activity. 5 Pretreatment of ureteral strips with the β‐adrenoceptor antagonist, propranolol (10−6 m), significantly increased the maximum contraction evoked by noradrenaline. After incubation with phentolamine (10−6 m), noradrenaline (10−7 − 10−6 m) decreased phasic activity induced by prostaglandin F2α (10−5 m). Isoprenaline and salbutamol also abolished PGF2α‐induced phasic activity. Pafenolol (10−6 m) and butoxamine (10−6 m) blocked the inhibitory effect of noradrenaline, isoprenaline, and salbutamol on PGF2α‐induced phasic activity. 6 These results suggest that noradrenaline may modulate both phasic peristaltic activity and basal tone of pig intravesical ureter through both α‐ and β‐adrenoceptors.

Prejunctional alpha sub 2-Adrenoceptors Inhibit Nitrergic Neurotransmission in Horse Penile Resistance Arteries

PURPOSE To study the influence of alpha-adrenergic stimuli on non-adrenergic non-cholinergic (NANC) neurogenic relaxation in isolated horse penile resistance arteries. MATERIALS AND METHODS Deep intracavernous penile arteries with an internal lumen diameter of 200-500 microns., isolated from the corpus cavernosum of young horses, were mounted in microvascular myographs for isometric tension recording and electrical field stimulation (EFS) of autonomic nerve terminals. RESULTS In the presence of guanethidine (10(-5) M) and atropine (10(-7) M) tone of the arteries was raised by the thromboxane analogue, U46619. EFS (1, 4 and 32 Hz) induced frequency-dependent relaxations, which were abolished in the presence of tetrodotoxin, while NG-nitro-L-arginine (L-NOARG, 10(-4) M) abolished the relaxations to EFS at 1 Hz, and significantly reduced the relaxations at 4 Hz and 32 Hz by 82.5 +/- 10.2% and 52.9 +/- 4.7%, respectively (n = 6). EFS induced relaxations of a similar magnitude in penile arteries contracted with U46619 or the alpha 1-adrenoceptor agonist, phenylephrine, while the alpha 2-adrenoceptor agonist, BHT920 (10(-6) M), produced an inhibitory effect on the EFS-evoked relaxations which was inversely related to the stimulus frequency (1, 4 and 32 Hz). BHT920 had no effect on the relaxations induced by exogenous nitric oxide (NO), added as acidified sodium nitrite (10(-6)-10(-3) M). The inhibitory effect of BHT920 on NANC relaxations was reversed by 10(-7) M rauwolscine. CONCLUSION These results suggest that the release of a NANC neurotransmitter primarily thought to be NO is inhibited by stimulation of prejunctional alpha 2-adrenoceptors in horse penile resistance arteries.

Involvement of nitric oxide in the non-adrenergic non-cholinergic neurotransmission of horse deep penile arteries: role of charybdotoxin-sensitive K(+)-channels.

1 . The involvement of nitric oxide (NO) and the signal transduction mechanisms mediating neurogenic relaxations were investigated in deep intracavernous penile arteries with an internal lumen diameter of 600–900 μm, isolated from the corpus cavernosum of young horses. 2 . The presence of nitric oxide synthase (NOS)‐positive nerves was examined in cross and longitudinal sections of isolated penile arteries processed for NADPH‐diaphorase (NADPH‐d) histochemistry. NADPH‐d‐positive nerve fibres were observed in the adventitia‐media junction of deep penile arteries and in relation to the trabecular smooth muscle. 3 . Electrical field stimulation (EFS) evoked frequency‐dependent relaxations of both endothelium‐intact and denuded arterial preparations treated with guanethidine (10−5 m) and atropine (10−7 m), and contracted with 10−6 m phenylephrine. These EFS‐induced relaxations were tetrodotoxin‐sensitive indicating their non‐adrenergic non‐cholinergic (NANC) neurogenic origin. 4 . EFS‐evoked relaxations were abolished at the lowest frequency (0.5‐2 Hz) and attenuated at higher frequencies (4–32 Hz) by the NOS inhibitor, NG‐nitro‐L‐arginine (L‐NOARG, 3 × 10−5m). This inhibitory effect was antagonized by the NO precursor, L‐arginine (3 × 10−3 m). NG‐nitro‐D‐arginine (10−4 m) did not affect the relaxations to EFS. 5 . Incubation with either the NO scavenger, oxyhaemoglobin (10−5 m), or methylene blue (10−5 m), an inhibitor of guanylate cyclase activation by NO, caused significant inhibitions of the EFS‐evoked relaxations, and while oxyhaemoglobin abolished the relaxations to exogenously added NO (acidified sodium nitrite, 10−6‐10−3m), there still persisted a relaxation to NO of 24.4 ± 5.1% (n = 6) in the presence of methylene blue. 6 . Glibenclamide (3 × 10−6 m), an inhibitor of ATP‐activated K+‐channels, did not alter the relaxations to either EFS‐stimulation or NO, while the blocker of Ca2+‐activated K+‐channels, charybdotoxin (3 × 10−8 m), caused a significant inhibition of both the electrically‐induced relaxations and the relaxations to exogenously added NO. Furthermore, charybdotoxin blocked relaxations induced by the cell permeable analogue of cyclic GMP, 8‐bromo cyclic GMP (8 Br‐cyclic GMP). 7 . These results suggest that relaxations of horse deep penile arteries induced by NANC nerve stimulation involve mainly NO or a NO‐like substance from nitrergic nerves. NO would stimulate the accumulation of cyclic GMP followed by increases in the open probability of Ca2+‐activated K+‐channels and hyperpolarization leading to relaxation of horse penile arteries.

Contribution of K+ channels and ouabain-sensitive mechanisms to the endothelium-dependent relaxations of horse penile small arteries

Penile small arteries (effective internal lumen diameter of 300–600 μm) were isolated from the horse corpus cavernosum and mounted in microvascular myographs in order to investigate the mechanisms underlying the endothelium‐dependent relaxations to acetylcholine (ACh) and bradykinin (BK). In arteries preconstricted with the thromboxane analogue U46619 (3–30 nM), ACh and BK elicited concentration‐dependent relaxations, pD2 and maximal responses being 7.71±0.09 and 91±1% (n=23), and 8.80±0.07 and 89±2% (n=24) for ACh and BK, respectively. These relaxations were abolished by mechanical endothelial cell removal, attenuated by the nitric oxide (NO) synthase (NOS) inhibitor, NG‐nitro‐L‐arginine (L‐NOARG, 100 μM) and unchanged by indomethacin (3 μM). However, raising extracellular K+ to concentrations of 20–30 mM significantly inhibited the ACh and BK relaxant responses to 63±4% (P<0.01, n=7) and to 59±4% (P<0.01, n=6), respectively. ACh‐ and BK‐elicited relaxations were abolished in arteries preconstricted with K+ in the presence of 100 μM L‐NOARG. In contrast to the inhibitor of ATP‐sensitive K+ channels, the blockers of Ca2+‐activated K+ (KCa) channels, charybdotoxin (30 nM) and apamin (0.3 μM), each induced slight but significant rightward shifts of the relaxations to ACh and BK without affecting the maximal responses. Combination of charybdotoxin and apamin did not cause further inhibition of the relaxations compared to either toxin alone. In the presence of L‐NOARG (100 μM), combined application of the two toxins resulted in the most effective inhibition of the relaxations to both ACh and BK. Thus, pD2 and maximal responses for ACh and BK were 7.65±0.08 and 98±1%, and 9.17±0.09 and 100±0%, respectively, in controls, and 5.87±0.09 (P<0.05, n=6) and 38±11% (P<0.05, n=6), and 8.09±0.14 (P<0.01, n=6) and 98±1% (n=6), respectively, after combined application of charybdotoxin plus apamin and L‐NOARG. The selective inhibitor of guanylate cyclase, 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ, 5 μM) did not alter the maximal responses to either ACh or BK, but slightly decreased the sensitivity to both agonists, δpD2 being 0.25±0.07 (P<0.05, n=6) and 0.62±0.12 (P<0.01, n=6) for ACh and BK, respectively. Combined application of ODQ and charybdotoxin plus apamin produced further inhibition of the sensitivity to both ACh (δpD2=1.39±0.09, P<0.01, n=6) and BK (1.29±0.11, P<0.01, n=6), compared to either ODQ or charybdotoxin plus apamin alone. Exogenous nitric oxide (NO) present in acidified solutions of sodium nitrite (NaNO2) and S‐nitroso‐cysteine (SNC) both concentration‐dependently relaxed penile resistance arteries, pD2 and maximal responses being 4.84±0.06 and 82±3% (n=12), and 6.72±0.07 and 85±4% (n=19), respectively. Charybdotoxin displaced to the right the dose‐relaxation curves for both NO (δpD2 0.38±0.06, P<0.01, n=6) and SNC (δpD2 0.50±0.10, P<0.01, n=5), whereas apamin only reduced sensitivity (δpD2=0.35±0.12, P<0.05, n=5) and maximum response (65±9%, P<0.05, n=6) to SNC. ODQ shifted to the right the dose‐relaxation curves to both NO and SNC. The relaxant responses to either NO or SNC were not further inhibited by a combination of ODQ and charybdotoxin or ODQ and charybdotoxin plus apamin, respectively, compared to either blocker alone. In the presence of 3 μM phentolamine, 5 μM ouabain contracted penile resistance arteries by 50±6% (n=17) of K‐PSS, but did not significantly change the relaxant responses to either ACh, BK or NO. However, in the presence of L‐NOARG ouabain reduced the ACh‐ and BK‐elicited relaxation from 94±3% to 16±5% (P<0.0001, n=6), and from 98±2% to 13±3% (P<0.0001, n=5), respectively. Combined application of ODQ and ouabain inhibited the relaxations to NO from 92±2% to 26±3% (P<0.0001, n=6). The present results demonstrate that the endothelium‐dependent relaxations of penile small arteries involve the release of NO and a non‐NO non‐prostanoid factor(s) which probably hyperpolarize(s) smooth muscle by two different mechanisms: an increased charybdotoxin and apamin‐sensitive K+ conductance and an activation of the Na+‐K+ATPase. These two mechanisms appear to be independent of guanylate cyclase stimulation, although NO itself can also activate charybdotoxin‐sensitive K+ channels and the Na+‐K+ pump through both cyclic GMP‐dependent and independent mechanisms, respectively.

Differential structural and functional changes in penile and coronary arteries from obese Zucker rats.

Erectile dysfunction frequently coexists with coronary artery disease and has been proposed as a potential marker for silent coronary artery disease in type 2 diabetes. In the present study, we comparatively assessed the structural and functional changes of both penile arteries (PAs) and coronary arteries (CAs) from a prediabetic animal model. PAs and CAs from 17- to 18-wk-old obese Zucker rats (OZRs) and from their control counterparts [lean Zucker rats (LZRs)] were mounted in microvascular myographs to evaluate vascular function, and stained arteries were subjected to morphometric analysis. Endothelial nitric oxide (NO) synthase (eNOS) protein expression was also assessed. The internal diameter was reduced and the wall-to-lumen ratio was increased in PAs from OZRs, but structure was preserved in CAs. ACh-elicited relaxations were severely impaired in PAs but not in CAs from OZRs, although eNOS expression was unaltered. Contractions to norepinephrine and 5-HT were significantly enhanced in both PAs and CAs, respectively, from OZRs. Blockade of NOS abolished endothelium-dependent relaxations in PAs and CAs and potentiated norepinephrine and 5-HT contractions in arteries from LZRs but not from OZRs. The vasodilator response to the phosphodiesterase 5 inhibitor sildenafil was reduced in both PAs and CAs from OZRs. Pretreatment with SOD reduced the enhanced vasoconstriction in both PAs and CAs from OZRs but did not restore ACh-induced relaxations in PAs. In conclusion, the present results demonstrate vascular inward remodeling in PAs and a differential impairment of endothelial relaxant responses in PAs and CAs from insulin-resistant OZRs. Enhanced superoxide production and reduced basal NO activity seem to underlie the augmented vasoconstriction in both PAs and CAs. The severity of the structural and functional abnormalities in PAs might anticipate the vascular dysfunction of the more preserved coronary vascular bed.

Adrenoceptor-mediated regulation of the contractility in horse penile resistance arteries.

The receptors mediating the contractions to both exogenously applied noradrenaline and electrical field stimulation (EFS) were characterized in horse isolated penile resistance arteries. The alpha 1-adrenoceptor-selective antagonist, prazosin, caused competitive rightward shifts of the contractile concentration-response curves (CRC) to phenylephrine. The alpha 2-antagonist, rauwolscine, also displaced to the right the CRC to the alpha 2-adrenoceptor-selective agonist, BHT 920. EFS (0.3 ms, 20-second trains) caused tetrodotoxin-sensitive frequency-dependent contractions which were enhanced in the presence of NG-nitro-L-arginine (L-NOARG, 3 x 10(-5) M), but not affected by mechanical endothelial cell removal. In experiments performed in the presence of L-NOARG, prazosin inhibited contractions to EFS, while rauwolscine inconsistently enhanced the contractile responses. Exogenously added noradrenaline induced contractions which were not changed in endothelium-denuded arteries, but significantly increased in the presence of L-NOARG. Prazosin inhibited the noradrenaline-induced contractions, while rauwolscine did not change the response to noradrenaline either alone or in the presence of prazosin. In the presence of phentolamine (10(-5) M), isoprenaline, adrenaline and the beta 2-adrenoceptor agonist, salbutamol, concentration-dependently relaxed penile resistance arteries, while the relaxations to noradrenaline and dobutamine, which activate beta 1-adrenoceptors, were negligible. Isoprenaline-induced relaxations were not changed in the presence of the beta 1-antagonist, atenolol (10(-7)-10(-6) M), but competitively inhibited by the beta 2-adrenoceptor antagonist, butoxamine (10(-6)-10(-5) M). The present results indicate that stimulation of adrenergic nerves in horse penile resistance arteries releases noradrenaline, which induces vasoconstriction through a predominant activation of alpha 1-adrenoceptors, while postjunctional alpha 2-adrenoceptors apparently play a minor role. Functional beta 2-adrenoceptors are also present in these arteries.

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.

Nitric oxide is involved in the non-adrenergic, non-cholinergic inhibitory neurotransmission of the pig intravesical ureter

NADPH-diaphorase histochemical staining and electrical field stimulation (EFS) were performed in vitro to investigate whether nitric oxide (NO) is involved in non-adrenergic non-cholinergic (NANC) inhibitory neurotransmission of pig intravesical ureter. NADPH-diaphorase activity was expressed in nerve trunks and thin nerve fibres around arteries and muscular bundles in the intravesical ureter. Relaxations to EFS were tetrodotoxin (10(-6) M)-sensitive which indicates their neurogenic origin. Addition of the NO-synthase inhibitor, L-NG-nitroarginine (L-NOARG, 3 x 10(-5) M), abolished the electrically induced relaxations, which were significantly reversed by L-arginine (3 x 10(-3) M). Addition of acidified sodium nitrite (NaNO2, 10(-5)-10(-3) M) evoked concentration-dependent relaxations of ureteral strips which were unaffected by L-NOARG. It is concluded that NO synthase is present in nerve fibres and NO seems to mediate the inhibitory neurotransmission of the porcine intravesical ureter.

Different muscarinic receptor subtypes mediating the phasic activity and basal tone of pig isolated intravesical ureter

1 We have studied the effects of muscarinic cholinoceptor agonists and specific antagonists on both phasic activity and basal tone of the isolated intravesical ureter of the pig by means of isometric techniques in vitro. 2 Acetylcholine in the presence and absence of physostigmine increased both phasic activity and basal tone of ureteral strips in a concentration‐dependent manner. Moreover carbachol, methacholine and oxotremorine‐M increased both contractile parameters while bethanechol and McN‐A‐343 evoked only increases in tone without affecting the frequency of the phasic contractions. 3 The nicotinic receptor blocker, hexamethonium (10−6–10−4 m), failed to modify the contractions evoked by a single dose of carbachol (10−5 m), whilst the muscarinic antagonist, atropine inhibited both phasic and tonic responses. 4 The muscarinic M1 (pirenzepine), M2 (AF‐DX 116 and methoctramine), M3 (4‐DAMP, HHSiD and p‐F‐HHSiD), and putative M4 receptor (tropicamide) antagonists significantly reversed increases in both frequency of phasic activity and baseline tone induced by a submaximal dose of carbachol (10−5 m). The pIC50 values for inhibition of the induced phasic activity were: atropine (10.16) > 4‐DAMP (9.12) > HHSiD (8.22) = methoctramine (7.98) = p‐F‐HHSiD (7.88 > tropicamide (7.62) = pirenzepine (7.53) = AF‐DX 116 (7.45) and for inhibition of basal tone were: atropine (10.73) > 4‐DAMP (9.32) > HHSiD (8.65) = pirenzepine (8.43) = p‐F‐HHSiD (8.38) > methoctramine (7.79) > tropicamide (7.53) > AF‐DX 116 (7.04). 5 The antagonist profile indicates that an M1 receptor mediates the tonic response while the phasic activity could involve either both M2 and M3 or an M4 muscarinic receptor. These results suggest that different muscarinic receptor subtypes mediate the phasic and tonic contractile activity induced by a submaximal concentration of carbachol in the porcine intravesical ureter.