Porfirio Carrillo

Electrophysiological evidence for the nomenclature of the pudendal nerve and sacral plexus in the male rat

Surgical microscopy and electrophysiological techniques were used to standardize the nomenclature for the pudendal nerve and sacral plexus according to their somatic axonal composition in the male rat. We conclude that the pudendal nerve is the segment running from the L6-S1 trunk to the sacral plexus, carrying efferent fibers to the coccygeus, internal obturator, ventral and dorsal bulbospongiosus, ischiocavernosus, external anal sphincter, and external urethral sphincter muscles, and afferent fibers from the penis, prepuce, scrotum, and ventral-proximal tail. The sacral plexus is the complex formed by the bridge-like structure connecting the pudendal nerve with the lumbosacral trunk, and two nerve branches emerging from it, one innervating the proximal half of the scrotal skin, and the other innervating the muscles at the base of the penis known as the motor branch. These branches are only considered as a part of the sacral plexus because they integrate axons from both the lumbosacral trunk and pudendal nerve. The gross anatomy of the pudendal nerve and sacral plexus has a main organization that was observed in 70% of cases, whereas the remaining 30% occurred in two variants. This nomenclature is appropriate to describe the pudendal nerve and sacral plexus in studies that involve them being lesioned or electrophysiologically analysed. A main additional finding was that two large afferent branches innervate the scrotum, one the proximal half and the other the distal half. As mentioned above, the proximal branch belongs to the sacral plexus, whereas the distal branch belongs to the pudendal nerve because all its axons travel to the cord via this nerve. Since stimulation or even manipulation of the scrotal branches resulted in the secretion of semen containing spermatozoa, it is suggested that scrotal afferents are involved in some way in the ejaculatory process, a topic that deserves further research.

Neurobiology of social attachments

Many types of social attachments can be observed in nature. We discuss the neurobiology of two types (1) intraspecific (with a partner) and (2) parental (with the offspring). Stimuli related to copulation facilitate the first, whereas pregnancy, parturition and lactation facilitate the second. Both types develop as consequence of cohabitation. These events seem to stimulate similar neural pathways that increase (1) social recognition, (2) motivation, reward; and (3) decrease fear/anxiety. Subregions of the amygdala and cortex facilitate social recognition and also disinhibition to decrease rejection responses. The interrelationship between MeA, BNST, LS may mediate the activation of NAcc via the mPOA to increase motivation and reward. Cortical areas such as the ACC discriminate between stimuli. The interaction between OT and D2-type receptors in NAcc shell facilitates intraspecific attachment, but D1-type appears to facilitate parental attachment. This difference may be important for maternal females to direct their attention, motivation and expression of attachment toward the appropriate target.

Fertility ratio in male rats: Effects after denervation of two pelvic floor muscles

Fertility ratio is defined here as the proportion of females that a male can impregnate after a constant period of in-polygyny living. This ratio was investigated in male rats after denervation of two pelvic floor muscles, the pubococcygeus and iliococcygeus. Denervation was carried out by transecting the somatomotor branch of the pelvic nerve. The lesion did not modify the sexual behavior of males or their overall fertility, but decreased the weight of the ejaculated seminal plug. Consequently, the number of days living in cohabitation to induce pregnancy was increased in lesioned males (approximately 13 days) compared with intact and sham animals (approximately 5 days). These results showed that the fertility ratio was optimal when intact/sham males cohabited with females for two consecutive estrous cycles, but that lesioned males needed up to four cycles to induce most pregnancies. Two hypotheses are raised by our results. The first is that pelvic floor denervation decreases the forceful tension required to expel the semen from the prostatic urethra to the vagina, then an incomplete seminal plug is expelled. The second is that denervation cut afferent fibers that reflexively promote the continence of the semen deposited in the prostatic urethra during seminal emission, allowing some to leak out before ejaculation. The latter hypothesis can also explain the recovery of the fertility ratio in lesioned males. It could be a compensatory mechanism mediated by the pudendal nerve supply to the coccygeus muscle, the other pelvic floor muscle.

The Role of Pubococcygeus Muscle in Urinary Continence in the Male Rat

The role of the male rat pubococcygeus muscle (Pcm) in the micturition reflex was analyzed. Anatomical features of Pcm, electrical stimulation of its nerve, electrical recording and stimulation of the muscle and cystometrograms were carried out. Results showed that Pcm has fibers attached to the ventrolateral part of the external urethral sphincter, and that its activity contributes to hold the tail in the midline. Pcm shows activity during fluid expulsion in cystometrograms and spontaneous micturitions. This activity produced reflex inhibition of detrusor contraction and was not the cause of intravesical high frequency oscillations. Thus, it is proposed that Pcm activity produces the discharge of its afferents which in turn activates a spinal reflex to promote continence.

Spinal organization and steroid sensitivity of motoneurons innervating the pubococcygeus muscle in the male rat.

Male rat motoneurons innervating the pubococcygeus muscle were located in the ventral nucleus of lamina IX at the sixth lumbar (L6) and first sacral (S1) spinal cord segments. Retrograde labeling with horseradish peroxidase‐wheat germ agglutinin was transported up to second‐order dendrites and revealed that these motoneurons have a “U‐shaped arborization” of dendrites toward the intermediolateral and intermediomedial nuclei area of lamina VII. This dendritic organization makes a wide “final common path” that probably integrates afferent information from several sources, accounting for the participation of the pubococcygeus muscle in autonomic and somatic processes, such as those related to micturition and reproduction. Castration produced a decrement in the morphometry of these motoneurons. A main effect was a decrement in dendritic length. Steroid replacement indicated that testosterone and estradiol, but not dihydrotestosterone, are able to induce a recovery of morphometric alterations. However, estrogen induced recovery after 2 weeks of treatment, whereas testosterone took 4 weeks. Thus, it is proposed that supraspinal aromatization of testosterone in the male central nervous system might be an important process for the appropriate organization of the pubococcygeus muscle motoneurons and that estradiol seems to need a shorter time of action than testosterone because of differential up‐regulation and down‐regulation of steroid receptors. J. Comp. Neurol. 409:358–368, 1999.

Effect of gonadal hormones on the cross-sectional area of pubococcygeus muscle fibers in male rat.

Effects of gonadal hormones on dimorphic striated muscles such as the bulbocavernosus/levator ani complex related to male penile erection have been widely studied. However, the action of these hormones on pelvic nondimorphic muscles is not known. In the present study, the sensitivity of the male rat pubococcygeus muscle (Pcm) to gonadal hormones was studied measuring the cross‐sectional area (CSA) of its fibers. For this, two experiments were done: in the first, the effect of castration, and in the second the effect of gonadal hormone administration was analyzed. We found that castration after 6 weeks significantly reduced the average CSA of the fibers of this muscle and that castration after 2 or 6 weeks reduced the percentage of fibers with higher CSAs, but only castration after 6 weeks increased the percentage of fibers with the lowest CSA. In comparison with castrated animals implanted with an empty Silastic capsule, Silastic implants of testosterone propionate or dihydrotestosterone significantly increased the average CSA of Pcm fibers, and the treatment with testosterone propionate, estradiol benzoate, or dihydrotestosterone decreased the percentage of fibers with low CSAs and increased the percentage with larger CSAs. Our results could be considered for therapy in patients with damage of the Pcm, and suffering urinary incontinence or ejaculatory dysfunctions. Anat Rec, 291:586–592, 2008.

Cutaneous Wounds Produced by Capsaicin Treatment of Newborn Rats Are Due to Trophic Disturbances

The purpose of this study was to show that the occurrence of skin ulcers observed in animals neonatally treated with the neurotoxin capsaicin coincide with trophic disturbances. In addition, cutaneous lesions increased when self-grooming and scratching behaviors reached maturity. The temporal course of cephalic cutaneous wounds in neonatally capsaicin-treated rats was evaluated in animals wearing and not wearing plastic collars from postnatal day (P) 21 until P45. The collars were used to prevent self-grooming and scratching. Beginning on P21, capsaicin-treated rats under both conditions showed transient skin ulcers distributed throughout the head and neck regions. In the capsaicin-treated group without collars, lesions reached their greatest severity by P40, when self-grooming and scratching behaviors obtained adult characteristics. Furthermore, no lesions were detected after 25 days. In the capsaicin-treated rats that wore plastic collars, the widest distribution of skin lesions occurred on P55, after which time lesions vanished detection by 25 days. In this latter group, the cutaneous lesions were exacerbated when collars were removed. Data suggest that transient cutaneous wounds associated with neonatal capsaicin administration may be mediated via capsaicin-sensitive sensory neurons that are involved in trophic and regenerating neural mechanisms.

Neuroendocrine control of urine-marking behavior in male rats

Sexually experienced Wistar male rats were used to investigate (a) urine voiding in the presence of nearby estrous females and the control of such voiding by (b) steroid hormones and (c) peripheral nerves supplying the genitourinary system. The first experiment showed that males always have a low rate of urine voiding that is significantly increased when a receptive female is around. Thus, it is suggested that an airborne scent from the female stimulates the olfactory system of males, triggering urine emission to transmit sex-related messages, i.e., male rats display the well-known urine-marking behavior of mammals. The number of urine marks and sniffing to females decreased after castration, and were restored after exogenous treatment with testosterone or estradiol. The proposed hypothesis is that airborne scents from the female activate the aromatization process in nuclei of the olfactory pathway of the male, evoking a cascade of neuronal responses that finish in urine marking. Peripheral nerves supplying the genitourinary system are the viscerocutaneous branch of the pelvic nerve (Vc) and the hypogastric (Hg). Data showed that both nerves are important for the central control of urine storage and voiding. Transection of Vc almost blocked urine marking, while Hg lesion increased the number of marks. Thus, it is discussed that Vc is the most important nerve in charge of voiding the bladder, and that Hg is important for continence.

Androgen receptors in Purkinje neurons are modulated by systemic testosterone and sexual training in a region-specific manner in the male rat

The androgen receptor (AR) is a widely distributed molecule indicating the spread actions of its ligand steroid, and plays an important role underlying male sexual behavior. Nevertheless, the influence of steroid hormones and their receptors on cerebellar neurons, as foundation of sexual behavior, is largely unknown. We sought to determine the influence of peripheral hormones on the AR expression in Purkinje neurons across cerebellar lobules in the vermis of male rats. First, we found a basal AR expression in Purkinje neurons that was higher in the superficial region than the deep region only in cerebellar lobules 2, 4, 5, 7, 8 and 9. Moreover, only the cerebellar lobule 10 showed a significant difference between the coordinates 0.1, 0.3 and 0.9. Second, males with four sessions of sexual training showed a decreased AR density in cerebellar lobules 7, 8, 9 and 10, but not in lobules 2, 4 or 5 when compared to males with one session of sexual training. However, sexual training did not affect AR expression in Purkinje neurons according to their location in any of the cerebellar lobules studied. Third, castration decreased the AR density in the cerebellar lobules 1, 2, 5 and 9 in the superficial region, while in the deep region all cerebellar lobules, except lobule 6, showed a lower AR density after castration. Finally, testosterone replacement restored AR density to control levels in all cerebellar lobules in the superficial region that were affected by castration. Contrary, in the deep region hormonal replacement failed to restore the AR density to control level in the majority of the cerebellar lobules that were affected by castration. Altogether, our findings indicate that Purkinje neurons in the vermis are influenced by systemic testosterone in a region-dependent manner highlighting a link between the cerebellum and gonads in the male rat. The AR function in Purkinje neurons may be related to cerebellar plasticity since both estrogen and progesterone receptors, members of the nuclear receptor family, regulate plasticity processes in Purkinje neurons. We concluded the cerebellum is an important component of the neural circuit for male sexual behavior.

Characteristics of ejaculated rat semen after lesion of scrotal nerves

The scrotum, representing the pouch surrounding the testes and their associated structures, plays a significant role in maintaining the gonad at a temperature lower than that of the body. Although thermoregulation of the testes has been ascribed as a main function of the scrotum, here we found that mechanical stimulation of the scrotum is important during mating to facilitate the appropriate expulsion of semen during ejaculation. Previously we showed that the scrotal skin area is innervated by two nerve branches, the proximal (Psb) and distal (Dsb) scrotal branches which supply the proximal or distal half of the scrotum, respectively. The sensory field of each nerve is testosterone-dependent. The decreased androgen levels following castration reduce the sensitive area to mechanical stimuli that can be restored following exogenous administration of the hormone. Here, we tested the effect of scrotal nerve transection on sexual parameters of experienced male rats. Data show that lesion of PSb or DSb alone or combined did not affect the execution of sexual behavior. However, these lesions significantly reduced the proportion of males that expelled semen during ejaculation, with that semen showing a reduced quantity of sperm. Thus, scrotal nerves are important in reproduction not for the appropriate display of sexual behavior, but for the expulsion of a normal quantity of semen and number of sperm during ejaculation. Our suggestion is that scrotal afferents trigger spinal reflexes to activate autonomic efferents supplying the male reproductive tract for the control of seminal emission.