Design and evaluation of nitrosylated alpha-adrenergic receptor antagonists as potential agents for the treatment of impotence.

We designed and evaluated a new class of molecules, nitrosylated alpha-adrenergic receptor antagonists, as potential agents for the treatment of impotence. In in vitro studies with human and rabbit corpus cavernosum strips in organ chambers, the alpha-adrenergic receptor antagonists (alpha-ARAs) moxisylyte and yohimbine and their corresponding nitrosylated compounds, SNO-moxisylyte (NMI-221) and SNO-yohimbine (NMI-187), concentration-dependently relaxed endothelin-induced contraction. The nitrosylated compounds were significantly more potent than the parent alpha-ARA. In human tissues, the specific phosphodiesterase type 5 inhibitor zaprinast potentiated the relaxing effects of the nitrosylated compounds. Only nitrosylated compounds induced accumulation of cyclic GMP in rabbit corpus cavernosum strips. Yohimbine and NMI-187 demonstrated a potent alpha2-blocking activity, with no significant differences in pA2 values (8.9 versus 8.2, respectively). Moxisylyte and NMI-221 showed moderate potency in antagonizing phenylephrine contraction, with comparable pA2 values for both molecules (6.5 versus 6.6, respectively). alpha-Adrenergic receptor-binding studies showed similar binding affinities for the alpha-ARA and their corresponding nitrosylated compounds. In vivo, intracavernosal injection of nitrosylated molecules caused greater increases in intracavernosal pressure (NMI-221 versus moxisylyte) that were more long lasting than those of moxisylyte or yohimbine. There were no significant differences between nitrosylated and non-nitrosylated compounds in the magnitude of systemic mean arterial pressure decrease after intracavernosal injection. alpha-ARA and the nitrosylated compounds showed no pain-inducing activity as evaluated with the paw-lick model in mice. In summary, nitrosylated alpha-ARA have the dual functionalities of nitric oxide donors and alpha-ARA. These drugs induced penile erection in animals, suggesting their possible therapeutic value as agents for the local pharmacological treatment of impotence.

Buckling of cylinders may cause prolonged penile pain after prosthesis implantation: a case control study using magnetic resonance imaging of the penis.

PURPOSE: We identify risk factors associated with chronic postoperative penile pain after inflatable prosthesis insertion. MATERIALS AND METHODS: We performed a case control study to determine the incidence of prolonged penile pain in 65 men who underwent prosthesis insertion. The association of various potential risk factors, including diabetes mellitus, incision site, prosthesis type, patient age and history of pelvic surgery, was assessed. In addition, the penis was examined for physical findings, and penile magnetic resonance imaging (MRI) was done as an in situ evaluation of the anatomical position and functional status of the inflatable prosthesis. RESULTS: A total of 14 patients (21.5%) with prolonged pain were compared to the remaining 51 with no pain. All parameters evaluated were similar in both groups except for MRI findings. Of the 14 patients with pain 12 (85.7%) had buckling of the cylinders in the flaccid state compared to only 1 of the 51 controls (1.9%). Statistical analysis showed a highly significant association between buckling and penile pain (odds ratio 300, p <0.0001). Surgical correction of buckling resolved pain in 5 patients. CONCLUSIONS: Prolonged postoperative penile pain after prosthesis insertion is strongly associated with cylinder buckling. This buckling may be the consequence of an excessively long cylinder or an appropriate size cylinder that fails to reach the crural end. The method of accurately diagnosing these alterations is MRI of the penis.

Engineering analysis of penile hemodynamic and structural-dynamic relationships: Part I--Clinical implications of penile tissue mechanical properties.

PURPOSE: The least investigated physical determinant of penile rigidity has been penile tissue material properties. The goals in this study (Part I) were to define two penile mechanical parameters, cavernosal expandability X and tunical distensibility VE/VF, determine their magnitudes in humans and develop an analytical expression for penile volume as a function of these two tissue characteristics and intracavernosal pressure. METHODS: Dynamic infusion pharmacocavernosometry was performed in 21 impotent patients (age 43 +/- 19 y) to provide human geometric, hemodynamic and structural data. A mathematically derived model of hemodynamic and structural-dynamic characteristics of penile erection was developed (Parts I, II, III) incorporating penile tissue mechanical qualities. RESULTS: Cavernosal expandability X provided a measure of the ability to approach maximum volume at relatively low intracavernosal pressures. Tunical distensibility VE/VF denoted the maximal erect to flaccid penile volume ratio. The magnitudes of X and VE/VF in the study population were 0.04-0.17 mmHg-1 and 1.7-5.0 respectively. CONCLUSIONS: Enabling penile volume to be derived as a function of tissue mechanical characteristics and pressure, allows for penile rigidity to be expressed (in Part II) as a function of pressure, geometry and tissue qualities.

Prostanoid production in rabbit corpus cavernosum: I. regulation by oxygen tension.

PURPOSE: To investigate the effects of oxygen tension on prostanoid synthesis in rabbit penile corpus cavernosum tissue (RCC) in organ culture. MATERIALS AND METHODS: Strips of rabbit corpus cavernosum were incubated in organ culture media under varying oxygen conditions (0%, 12% and 21% oxygen), in the presence or absence of acetylcholine and arachidonate stimulation. Prostanoids were measured in collected media by radioimmunoassay. Prostaglandin H synthase (PGHS) protein levels and mRNA PGHS expression were measured under both 0% and 21% oxygen conditions. RESULTS: Basal and acetylcholine-stimulated PGI2 release was progressively diminished as a function of diminishing oxygen tension (pO2 from approximately 165 to 25 mm.Hg). The basal and stimulated production of other prostanoids, thromboxane A2, PGF2alpha, and PGE2, was also significantly inhibited under 0% oxygen (approximately 25 mm.Hg) conditions. However, incubation under 0% oxygen did not alter PGHS protein levels nor mRNA PGHS expression. Cavernosal strips incubated under 0% oxygen but supplemented with exogenous arachidonate (10 microM.) maintained significantly lower PGI2 production than tissues exposed to 21% oxygen (approximately 165 mm.Hg). CONCLUSIONS: These data demonstrate that oxygen tension regulates prostaglandin production in corporal tissue. The reduction in prostanoid production during hypoxia can be attributed to inhibition of PGHS activity rather than the expression of the enzyme. In view of the role of PGI2 as an inhibitor of platelet aggregation and white cell-endothelial adhesion, our findings may provide mechanistic insight into the alteration in corporal blood homeostasis ischemic-hypoxic priapism.

Identification of alpha 1-adrenergic receptor subtypes in human corpus cavernosum tissue and in cultured trabecular smooth muscle cells.

Recent pharmacological and functional studies have suggested the presence of more than one alpha-1 adrenergic receptor subtype in human corpus cavernosum (HCC). In this study, we sought to identify the alpha-1 adrenergic receptor (alpha 1-AR) subtypes expressed in HCC whole tissue and in trabecular smooth muscle subcultured from this tissue. We have utilized RNase protection assays and in situ hybridization (ISH) techniques to identify and localize these receptor subtypes. RNase protection assays of mRNA isolated from whole tissue demonstrated the presence of mRNA transcripts for three alpha 1-AR receptor subtypes (alpha 1d, alpha 1b, and alpha 1a). alpha 1d-AR and alpha 1a-AR appear to be more abundant than alpha 1b-AR. The identification and localization of mRNA for alpha 1-AR subtypes in whole tissue was demonstrated by RNA protection assays and ISH analysis. Immunocytochemical analysis of alpha 1-AR by an antipeptide antibody developed against a specific amino acid sequence derived from alpha 1d-AR subtype demonstrated specific staining of the smooth muscle cells, suggesting the expression of alpha 1d-AR subtype. In cultured HCC smooth muscle cells (HCC SMC), phenylephrine,alpha 1-AR agonist stimulated Na+/K+ ATPase activity, suggesting the presence of functional alpha 1-AR. RNase protection assay of mRNA isolated from HCC SMC grown in culture further demonstrated the presence of mRNA transcripts for alpha 1d-AR and alpha 1a-AR subtypes. ISH analysis and confocal microscopy also indicate that the SMC express the alpha 1d-AR and alpha 1a-AR subtypes. The data presented suggests that HCC and SMC derived from this tissue express at least three alpha 1-AR subtypes. Identification of these receptor subtypes should allow characterization of the functional role of these receptor subtypes in regulation of trabecular smooth muscle tone and penile detumescence.

Characterization of muscarinic acetylcholine receptors in cultured bovine aortic endothelial cells.

The binding characteristics of [3H]quinuclidinyl benzilate ([3H]QNB) to isolated crude membranes of cultured bovine aortic endothelial cells were investigated. [3H]QNB bound to endothelial cell membranes with high affinity (kD = 0.056 nM) and limited capacity (132 fmol/mg DNA). The binding specificity, order of affinity and inhibition constants (Ki) were determined by displacement of bound [3H]QNB with unlabeled ligands. The order of affinity was QNB > atropine > 4-diphenylacetoxy-N-methyl-piperidine methiodide (4-DAMP) > p-fluoro-hexahydro-sila-difenidol (p-F-HHSiD) (M3 antagonist) > pirenzepine (M1 antagonist) > AFDX-116 (M2 antagonist) > (4-hydroxy-2-butynyl) trimethylammonium chloride m-chlorocarbanilate (McN-A-343, M1 agonist). These observations suggest that muscarinic receptors of endothelial cells in culture are likely to be of M3 and M1 subtype. Northern blot analysis of receptor subtypes using cDNA probes did not provide conclusive results due to the low level expression of these receptors in cultured cells. Solubilization of protein bound [3H]QNB with 1% digitonin and 0.02% cholate followed by analysis on sucrose density gradients demonstrated the presence of a specifically bound [3H]QNB-protein complex sedimenting at the 6.2S region of the gradient. These data demonstrate the presence of muscarinic acetylcholine receptor protein in cultured bovine aortic endothelial cells.