F Giuliano

The efficacy and tolerability of vardenafil, a new, oral, selective phosphodiesterase type 5 inhibitor, in patients with erectile dysfunction: the first at-home clinical trial.

Vardenafil, a novel selective phosphodiesterase type 5 inhibitor, was evaluated in its first large-scale at-home trial. A total of 601 men with mild to severe erectile dysfunction (ED) were enrolled in this multi-centre, randomized, double-blind, placebo-controlled trial of 12 weeks of treatment with either placebo or 5, 10 and 20u2005mg of vardenafil. Primary endpoints were Q3 (vaginal penetration) and Q4 (maintenance of erection) of the International Index of Erectile Function (IIEF). In the intent-to-treat population (n=580), the changes from baseline for 5, 10 and 20u2005mg vardenafil (1.2, 1.3 and 1.5, respectively) were all improved (P<0.001) over placebo (0.2) for Q3 and were similarly improved for Q4 (1.4, 1.5 and 1.7) compared to placebo (0.5) (P<0.001). All vardenafil doses improved all IIEF domains compared to placebo (P<0.001). The percentage of successful intercourses was between 71 and 75% for the three vardenafil doses. For the 20u2005mg dose, 80% of the patients experienced improved erections (GAQ) compared to 30% for placebo. Most frequent treatment-emergent adverse events were headache (7–15%), flushing (10–11%) and up to 7% for dyspepsia or rhinitis. Vardenafil treatment resulted in a high efficacy and low adverse-event profile in a population with mixed ED etiologies.

Oxytocinergic innervation of autonomic nuclei controlling penile erection in the rat

In the rat, spinal autonomic neurons controlling penile erection receive descending pathways that modulate their activity. The paraventricular nucleus of the hypothalamus contributes oxytocinergic fibers to the dorsal horn and preganglionic sympathetic and parasympathetic cell columns. We used retrograde tracing techniques with pseudorabies virus combined with immunohistochemistry against oxytocin and radioligand binding detection of oxytocinergic receptors to evidence the oxytocinergic innervation of thoracolumbar and lumbosacral spinal neurons controlling penile erection. Spinal neurons labelled with pseudo-rabies virus transsynaptically transported from the corpus cavernosum were present in the intermediolateral cell column and the dorsal gray commissure of the thoracolumbar and lumbosacral spinal cord. Confocal laser scanning microscopic observation of the same preparations revealed close appositions between oxytocinergic varicosities and pseudorabies virus-infected neurons, suggesting strongly the presence of synaptic contacts. Electron microscopy confirmed this hypothesis. Oxytocin binding sites were present in the superficial layers of the dorsal horn, the dorsal gray commissure and the intermediolateral cell column in both the thoracolumbar and lumbosacral segments. In rats, stimulation of the paraventricular nucleus induces penile erection, but the link between the nucleus and penile innervation remains unknown. Our findings support the hypothesis that oxytocin, released by descending paraventriculo-spinal pathways, activates proerectile spinal neurons.

Oxytocinergic and serotonergic innervation of identified lumbosacral nuclei controlling penile erection in the male rat

Penile erection is due to activation of proerectile neurons located in the sacral parasympathetic nucleus of the L6-S1 spinal cord in the rat. Contraction of the ischiocavernosus and bulbospongiosus striated muscles, controlled by motoneurons located in the ventral horn of the L5-L6 spinal cord, reinforces penile erection. Physiological and pharmacological arguments have been provided for a role of oxytocin and serotonin in the spinal regulation of penile erection. Immunohistochemistry of oxytocinergic and serotonergic fibres was performed at the lumbosacral level of the male rat spinal cord, and combined with retrograde tracing from the pelvic nerve or from the ischiocavernosus and bulbospongiosus muscles using wheat germ agglutinin-horseradish peroxidase. Sacral preganglionic neurons retrogradely labelled from the pelvic nerve formed a homogeneous population, predominant at the L6 level. Motoneurons retrogradely labelled from the ischiocavernosus and bulbospongiosus muscles were observed in the medial part of the dorsolateral and in the dorsomedial nuclei. Fibres immunoreactive for oxytocin were mainly distributed in the superficial layers of the dorsal horn, the dorsal gray commissure and the sacral parasympathetic nucleus. Some of these fibres were apposed to retrogradely-labelled sacral preganglionic neurons and at the ultrastructural level, some synapses were evidenced. Fibres immunoreactive for serotonin were largely and densely distributed in the dorsal horn, the dorsal gray commissure, the sacral parasympathetic nucleus and the ventral horn. Some serotonergic fibres occurred in close apposition with retrogradely-labelled sacral preganglionic neurons and motoneurons, and synapses were demonstrated at the ultrastructural level. This study provides morphological support for a role of oxytocin and serotonin on sacral preganglionic neurons innervating pelvic organs and motoneurons innervating the ischiocavernosus and bulbospongiosus muscles.

5-Hydroxytryptamine2C receptors on spinal neurons controlling penile erection in the rat

The localization of 5-hydroxytryptamine2C receptors in the lumbosacral spinal cord of the rat was investigated using selective antibodies raised against the carboxyl-terminal part of the rat receptor. The distribution of immunoperoxidase labelling at the light microscope level revealed numerous labelled neurons in the gray matter, with a higher intensity in the sacral parasympathetic nucleus, the dorsal gray commissure and particularly the motoneurons of the ventral horn. Confocal microscope analysis showed that immunostaining was mainly intracellular (motoneurons), but could also be associated with the membrane of cell bodies and dendrites. Actually, electron microscope immunogold experiments demonstrated an exclusive staining of the cis-Golgi apparatus. Following pseudo-rabies virus transsynaptic retrograde labelling from the corpus cavernosum, labelled neurons were found in the sacral parasympathetic nucleus and the dorsal gray commissure of the L6-S1 segments. All virus-labelled neurons exhibited 5-hydroxytryptamine2C receptor immunoreactivity. These results indicate that all parasympathetic preganglionic neurons and their related interneurons which contribute to the innervation of cavernosal tissue bear 5-hydroxytryptamine2C receptors. In the sacral parasympathetic nucleus, most neurons which were retrogradely-labelled from the pelvic ganglion with Fast Blue also showed 5-hydroxytryptamine2C receptor immunoreactivity. In the ventral horn, motoneurons retrogradely labelled from the ischiocavernosus muscle and the bulbospongiosus muscle, both of which are involved in erection and ejaculation, were also 5-hydroxytryptamine2C receptor-immunopositive. The supraspinal serotoninergic control of erection at the lumbosacral level therefore appears to be strongly associated with the activation of 5-hydroxytryptamine2C receptors, consistent with the proerectile properties of 5-hydroxytryptamine2C agonists.

The 'effectiveness' scale--therapeutic outcome of pharmacologic therapies for ED: an international consensus panel report.

Despite availability of outcome measures and scales for assessing erectile dysfunction (ED) treatment efficacy, guidelines are not available for assessing broader therapeutic outcomes or defining treatment failure in ED. An International Consensus Advisory Panel was convened to develop guidelines, definitions and a new algorithm for evaluating treatment effectiveness in ED. These new guidelines are recommended for use in both research and clinical practice. A multidisciplinary, international panel, consisting of 11 senior researchers and clinicians, was convened to address pertinent issues concerning therapeutic outcome assessment for ED. The panel utilized a modified Delphi method of consensus development and proposed a new model for outcomes assessment. This model is inherently testable, using existing instruments and current methods of assessment. Following a comprehensive literature review and discussion, the Panel recommended adoption of a new treatment effectiveness conceptual framework or theoretical model for assessing therapeutic outcomes in ED. Treatment effectiveness is presumed to be a combined function of two other factors, treatment response and treatment satisfaction. Treatment response is based on the combined assessment of efficacy and tolerability, and treatment satisfaction on the combined assessment of patient and partner satisfaction. Taken together, these two domains define an overall domain of treatment effectiveness. This therapeutic index would be derived by independently assessing treatment efficacy and satisfaction by means of event logs, questionnaires or the more typical patient interview methods. In conclusion, the Ad Hoc Advisory Consensus Panel recommends adoption of a new framework or conceptual model for conducting ED outcome trials or clinical research. The concept of ‘treatment effectiveness’ is proposed as a new ‘umbrella concept’ or distal outcome to be evaluated.

Identification of lumbar spinal neurons controlling simultaneously the prostate and the bulbospongiosus muscles in the rat.

Lumbar spinothalamic neurons in the lamina X of the L3-L4 spinal cord segment have been proposed to constitute the spinal ejaculation generator in male rats. Lumbar spinothalamic cells are immunoreactive for galanin and neurokinin-1 receptors. We previously showed that after injection of pseudorabies virus either in the bulbospongiosus muscle or in the prostate, retrogradely labeled cells in the L3-L4 segment also displayed galanin or neurokinin-1 receptor immunoreactivities, demonstrating a direct link between lumbar spinothalamic cells and two anatomical structures involved in the two phases of ejaculation i.e. the emission and the expulsion phases. In order to provide with a more precise anatomical support for the role of lumbar spinothalamic cells in controlling ejaculation, we injected simultaneously in male adult rats two strains of recombinant pseudorabies virus, expressing either beta-galactosidase (PRV-BaBlu) or green fluorescent protein (PRV-152) in the prostate and in the bulbospongiosus muscle, respectively. After 5 days, we performed multiple immunofluorescence experiments to detect PRV-BaBlu, PRV-152 and galanin or neurokinin-1 receptors in transverse sections of the L1-S1 segment. Double- and triple-labeled cells were counted using confocal laser scanning microscope. Double-labeled neurons with the two strains of pseudorabies virus were mainly found at the L3-L4 segment lateral to the central canal in lamina X and represented about 60% of the total number of pseudorabies virus-labeled neurons. All the double pseudorabies virus-labeled neurons also expressed lumbar spinothalamic and most of them neurokinin-1 receptor, identifying them as lumbar spinothalamic neurons. The convergence of retrograde labeling from prostate and bulbospongiosus muscle on the same lumbar spinothalamic cells strongly reinforce their role in the spinal control and coordination of the emission and expulsion of sperm.

Telemetric Monitoring of Intracavernous Pressure in Freely Moving Rats During Copulation

Changes in intracavernous pressure (ICP) were recorded in freely moving rats by means of a catheter implanted in the corpus cavernosum and connected to a pressure transducer located subcutaneously. Intracavernous pressure changes and copulatory events (mounts, intromissions and ejaculations) were recorded during several overnight testing sessions with 12 rats in the presence of receptive females. Video recordings of sexual behavior in the implanted and nonimplanted rats revealed that the ICP device did not impair copulatory behavior. Comparison between copulatory behavior and ICP changes revealed that ICP profiles could identify the number of intromissions and ejaculations as well as the timing of these events. The results showed that during mounts, intromissions and ejaculations, ICP increases were 36.9 mm. Hg, 62 mm. Hg and 106.3 mm. Hg in 4 rats. The increases in ICP during mounts and intromissions appeared higher in sexually experienced as compared with sexually naive rats. These results demonstrate the reliability of the telemetric measurement of ICP in freely moving rats under natural physiological conditions. The model provides quantitative data for research on the neurophysiology of penile erection and evaluation of oral treatments for impotence.

Galanin and neurokinin-1 receptor immunoreactivity spinal neurons controlling the prostate and the bulbospongiosus muscle identified by transsynaptic labeling in the rat

Ejaculation requires the coordination of sympathetic, parasympathetic and somatic neural outputs. Timely occurrence of the emission and expulsion of sperm results from an interplay between spinal nuclei innervating the seminal tract and the sexual accessory glands including the prostate on the one hand, and on the other hand perineal striated muscles, particularly the bulbospongiosus muscle. A group of cells essential for ejaculation, located around the central canal and referred to as lumbar spinothalamic neurons have been recently identified. Lumbar spinothalamic neurons are immunoreactive for galanin and neurokinin-1 receptor. In order to investigate the anatomical relationships between lumbar spinothalamic neurons and both the prostate and the bulbospongiosus muscle, pseudorabies virus retrograde tracing technique was used combined with immunohistochemistry. Three to five days after pseudorabies virus injection in the bulbospongiosus muscle or the prostate in male rats, spinal cord sections were processed for double immunofluorescence against pseudorabies virus and galanin or neurokinin-1 receptor. Immunocytochemical experiments against pseudorabies virus and choline acetyltransferase were also performed to discriminate between motoneurons and preganglionic neurons, or interneurons. Spinal sections were examined with confocal laser scanning microscope. Three days after pseudorabies virus injection within the prostate and the bulbospongiosus muscle, sympathetic preganglionic neurons and motoneurons of the dorsomedial nucleus were retrogradely labeled, respectively. Five days after pseudorabies virus injection, transsynaptically labeled choline acetyltransferase-negative neurons were found mainly located in the medial gray surrounding the central canal from L1 to S1. At the L3-L4 level, most of transsynaptically labeled neurons were immunoreactive for galanin and to a lesser extent for neurokinin-1 receptor, strongly suggesting that they could be the lumbar spinothalamic cells. We have thus evidenced connections between these cells and motoneurons of the dorsomedial nucleus and both sympathetic and parasympathetic preganglionic neurons innervating the bulbospongiosus muscle and the prostate, respectively. These anatomical data reinforce the crucial role for lumbar spinothalamic cells in coordinating the spinal control of ejaculation.

Spinal proerectile effect of apomorphine in the anesthetized rat

Considering the presence of dopaminergic receptors in the lumbosacral spinal cord, we tested whether apomorphine could exert a proerectile effect by acting at the spinal level. Intracavernous (ICP) and blood pressures (BP) were measured in anesthetized rats. ICP rises were quantified (duration, percentage of ICPmaximum/meanBP (ICPmax/BP×100), area under ICP curve (AUC/BP) and sum of AUC/BP after intravenous (i.v.) and intrathecal (i.t.) injections of apomorphine alone or in presence of i.t. oxytocin (10 ng). Both 10 and 30 µg i.v. apomorphine dosings elicited erectile events evidenced by ICP rises. Upon the 30 µg i.v. injection, duration of ICP rises were increased from 25±10 to 69±18 s (P<0.001), ICPmax/BP×100 from 21±3 to 50±14% (P=0.001), AUC/BP from 3±1 to 14±6 s (P=0.002) and sum of AUC/BP from 5±7 to 34±35 s (P=0.021). Upon 30 µg i.t. injections of apomorphine at the lumbosacral level, the number of ICP rises was increased from 0.2±0.4 to 3.0±1.5, ICPmax/BP×100 from 16±9 to 43±12 and sum of AUC/BP from 1±3 to 31±15 s compared to vehicle injection (P<0.05 for all parameters). Injection of 30 µg i.v. or i.t. apomorphine non-significantly enhanced the number and amplitude of the ICP rises induced by 10 ng i.t. oxytocin. However, the enhancement of the amplitude of the ICP rises elicited by i.t. oxytocin was more pronounced with i.t. apomorphine than with i.v. apomorphine. These results suggest the existence of a spinal site of action for apomorphine which may (1) participate to generation of erection and (2) exerts a facilitator effect on erection of supraspinal origin.

Electrophysiological Study of Relations between the Dorsal Nerve of the Penis and the Lumbar Sympathetic Chain in the Rat

Afferent sensory inputs from the penis are carried by the dorsal nerve of the penis (DNP) to the spinal cord. Sympathetic outflow involved in the control of the urogenital tract is partly conveyed by the lumbosacral sympathetic chain. Our aim was to search for a sympathetic component in the DNP and relations between DNP afferents and sympathetic fibers conveyed by the distal sympathetic chain in anesthetized adult male rats. Stimulation of the lumbar sympathetic chain at the L4-L5 level (LSC4-5) elicited an evoked discharge on the DNP. This discharge was abolished by cutting the sympathetic chain distal to the stimulation site. Ganglionic blockade with hexamethonium and various neural sections revealed the presence of sympathetic postganglionic fibers in the DNP, originating in the sympathetic chain. Stimulation of the DNP evoked a reflex discharge in the LSC. This reflex was spinally mediated since it was abolished by acute spinal cord transection at the L5 level. Acute spinalization at the T8 level significantly reduced the latency of the evoked response. We hypothesize that both spinal and supraspinal control exist over relations of the DNP afferents with sympathetic outflow to the pelvis. Increase in sympathetic tone elicited by activation of penile sensory fibers could play a role in regulation of sexual function.