Excitotoxic Lesions of the Nucleus Paragigantocellularis Facilitate Male Sexual Behavior but Attenuate Female Sexual Behavior in Rats

Little is known regarding the descending inhibitory control of genital reflexes such as ejaculation and vaginal contractions. The brainstem nucleus paragigantocellularis (nPGi) projects bilaterally to the lumbosacral motoneuron pools that innervate the genital musculature of both male and female rats. Electrolytic nPGi lesions facilitate ejaculation in males, leading to the hypothesis that the nPGi is the source of descending inhibition to genital reflexes. However, the function of the nPGi in female sexual behavior remains to be elucidated. To this end, male and female rats received bilateral excitotoxic fiber-sparing lesions of the nPGi, and sexual behavior and sexual behavior-induced Fos expression were examined. In males, nPGi lesions facilitated copulation, supporting the hypothesis that the nPGi, and not fibers-of-passage, is the source of descending inhibition of genital reflexes in male rats. nPGi lesions in males did not alter sexual behavior-induced Fos expression in any brain region examined. nPGi-lesioned females spent significantly less time mating with stimulus males and had significantly longer ejaculation-return latencies compared to baseline. These results did not significantly differ from control females, but this trend warranted further analysis of the reinforcing value of sexual behavior. Both lesioned and non-lesioned females formed a conditioned place preference (CPP) for artificial vaginocervical stimulation (aVCS). However, post-reinforcement, nPGilesioned females did not differ in the percentage of time in spent in the non-reinforced chamber versus the reinforced chamber, suggesting a weakened CPP for aVCS. nPGi lesions in females reduced sexual behavior-induced Fos expression throughout the hypothalamus and amygdala. Taken together, these results suggest that while nPGi lesions in males facilitate copulation, such lesions in females attenuate several aspects of sexual behavior resulting in a reduction in the rewarding value of copulation that may be mediated by nPGi control of genital reflexes. This work has important implications for the understanding and treatment of sexual dysfunction in people including delayed/premature ejaculation, involuntary vaginal spasms, and pain during intercourse

Sex Differences in the Anatomical and Functional Organization of the Periaqueductal Gray-Rostral Ventromedial Medullary Pathway in the Rat: A Potential Circuit Mediating the Sexually Dimorphic Actions of Morphine

Previous studies have demonstrated that morphine, administered systemically or directly into the PAG, produces a significantly greater degree of antinociception in males in comparison to females. As the midbrain periaqueductal gray (PAG) and its descending projections to the rostral ventromedial medulla (RVM) constitute an essential neural circuit for opioid-based analgesia, the present studies were conducted to determine if sex differences in the anatomical organization of the PAG-RVM pathway, and its activation during persistent inflammatory pain, could account for sex-based differences in opioid analgesia. In the rat, retrograde tracing was combined with Fos immunocytochemistry to investigate sexual dimorphism in the organization of the PAGRVM circuit and its activation by persistent inflammatory pain induced by intraplantar injection of complete Freund’s adjuvant (CFA). The ability of morphine to suppress the activation of the PAG-RVM circuit was also examined. Sexually dimorphic retrograde labeling was observed within the dorsomedial and lateral/ventrolateral PAG at all rostrocaudal levels, with females having significantly more PAG-RVM output neurons in comparison to males. While no sex differences were noted in the activation of the PAGRVM circuit by persistent inflammatory pain, significantly more double labeled cells were found in males in comparison to females. Systemic administration of morphine significantly suppressed CFA-induced Fos in the PAG in males only. The results of these studies demonstrate that both the anatomical organization, and functional activation, of the PAG-RVM circuit is sexually dimorphic, and may provide the anatomical substrate for sex-based differences in morphine analgesia

Neonatal Injury Alters Adult Pain Sensitivity by Increasing Opioid Tone in the Periaqueductal Gray

Studies in both rodents and humans have shown that acute inflammatory pain experienced during the perinatal period produces long-term decreases in pain sensitivity (hypoalgesia) (Grunau et al., 1994a , 2001 ; Ren et al., 2004 ; LaPrairie and Murphy, 2007 ). To date, the mechanisms underlying these long-term adaptations, however, have yet to be elucidated. The present studies tested the hypothesis that neonatal inflammatory pain induces an upregulation in endogenous opioid tone that is maintained into adulthood, and that this increase in opioid tone provides the underlying mechanism for the observed hypoalgesia. On the day of birth (P0), inflammatory pain was induced in male and female Sprague-Dawley rats by intraplantar administration of carrageenan (CGN; 1%). In adulthood (P60), these animals displayed significantly increased paw withdrawal latencies in response to a noxious thermal stimulus in comparison to controls. Systemic administration of the brain-penetrant opioid receptor antagonist naloxone HCl, but not the peripherally restricted naloxone methiodide, significantly attenuated the injury-induced hypoalgesia. Direct administration of naloxone HCl or antagonists directed at the mu or delta opioid receptors into the midbrain periaqueductal gray (PAG) also significantly reversed the injury-induced hypoalgesia in adult rats. Parallel anatomical studies revealed that inflammatory pain experienced on the day of birth significantly increased beta-endorphin and met/leu-enkephalin protein levels and decreased opioid receptor expression in the PAG of the adult rat. Thus, early noxious insult produces long-lasting alterations in endogenous opioid tone, thereby profoundly impacting nociceptive responsiveness in adulthood

Long Term Impact of Neonatal Injury in Male and Female Rats: Sex Differences, Mechanisms and Clinical Implications

Over the last several decades, the relative contribution of early life events to individual disease susceptibility has been explored extensively. Only fairly recently, however, has it become evident that abnormal or excessive nociceptive activity experienced during the perinatal period may permanently alter the normal development of the CNS and influence future responses to somatosensory input. Given the significant rise in the number of premature infants receiving high‐technology intensive care over the last twenty years, ex‐preterm neonates may be exceedingly vulnerable to the long‐term effects of repeated invasive interventions. The present review summarizes available clinical and laboratory findings on the lasting impact of exposure to noxious stimulation during early development, with a focus on the structural and functional alterations in nociceptive circuits, and its sexually dimorphic impact

Persistent Peripheral Inflammation Attenuates Morphine-induced Periaqueductal Gray Glial Cell Activation and Analgesic Tolerance in the Male Rat

Morphine is among the most prevalent analgesics prescribed for chronic pain. However, prolonged morphine treatment results in the development of analgesic tolerance. An abundance of evidence has accumulated indicating that CNS glial cell activity facilitates pain transmission and opposes morphine analgesia. While the midbrain ventrolateral periaqueductal gray (vlPAG) is an important neural substrate mediating pain modulation and the development of morphine tolerance, no studies have directly assessed the role of PAG‐glia. Here we test the hypothesis that morphine‐induced increases in vlPAG glial cell activity contribute to the development of morphine tolerance. As morphine is primarily consumed for the alleviation of severe pain, the influence of persistent inflammatory pain was also assessed. Administration of morphine, in the absence of persistent inflammatory pain, resulted in the rapid development of morphine tolerance and was accompanied by a significant increase in vlPAG glial activation. In contrast, persistent inflammatory hyperalgesia, induced by intraplantar administration of Complete Freund’s Adjuvant (CFA), significantly attenuated the development of morphine tolerance. No significant differences were noted in vlPAG glial cell activation for CFA‐treated animals versus controls. These results indicate that vlPAG glia are modulated by a persistent pain state, and implicate vlPAG glial cells as possible regulators of morphine tolerance. The development of morphine tolerance represents a significant impediment to its use in the management of chronic pain. We report that morphine tolerance is accompanied by increased glial cell activation within the vlPAG, and that the presence of a persistent pain state prevented vlPAG glial activation and attenuated morphine tolerance

Serotonergic Lesions of the Periaqueductal Gray, a Primary Source of Serotonin to the Nucleus Paragigantocellularis, Facilitate Sexual Behavior in Male Rats

While selective serotonin reuptake inhibitors (SSRIs) are widely used to treat anxiety and depression, they also produce profound disruptions of sexual function including delayed orgasm/ejaculation. The nucleus paragigantocellularis (nPGi), a primary source of inhibition of ejaculation in male rats, contains receptors for serotonin (5-HT). The ventrolateral periaqueductal gray (vlPAG) provides serotonin to this region, thus providing an anatomical and neurochemical basis for serotonergic regulation of the nPGi. We hypothesize that 5-HT acting at the nPGi could underlie the SSRI-induced inhibition of ejaculation in rodents. To this end, we produced 5-HT lesions of the source of 5-HT to the nPGi (the vlPAG) and examined sexual behavior. Removing the source of 5-HT to the nPGi facilitated genital reflexes, but not other aspects of sexual behavior, consistent with our hypothesis. Namely, 5-HT lesions produced a significant increase in the mean number of ejaculations and a significant decrease in ejaculation latency as compared to sham lesioned animals, while latency to mating and the post-ejaculatory interval did not differ. These data suggest that the serotonergic vlPAG-nPGi pathway is an important regulatory mechanism for the inhibition of ejaculation in rats, and supports the hypothesis that this circuit contributes to SSRI-induced inhibition of ejaculation

Somatic Genital Reflexes in Rats with a Nod to Humans: Anatomy, Physiology, and the Role of the Social Neuropeptides

Somatic genital reflexes such as ejaculation and vaginocervical contractions are produced through the striated muscles associated with the genitalia. The coordination of these reflexes is surprisingly complex and involves a number of lumbosacral spinal and supraspinal systems. The rat model has proved to be an excellent source of information regarding these mechanisms, and many parallels to research in humans can be drawn. An understanding of the spinal systems involving the lumbosacral spinal cord, both efferent and afferent, has been generated through decades of research. Spinal and supraspinal mechanisms of descending excitation, through a spinal ejaculation generator in the lumbar spinal cord and thalamus, and descending inhibition, through the ventrolateral medulla, have been identified and characterized both anatomically and physiologically. In addition, delineation of the neural circuits whereby ascending genitosensory information regarding the regulation of somatic genital reflexes is relayed supraspinally has also been the topic of recent investigation. Lastly, the importance of the “social neuropeptides” oxytocin and vasopressin in the regulation of somatic genital reflexes, and associated sociosexual behaviors, is emerging. This work not only has implications for understanding how nervous systems generate sexual behavior, but also provides treatment targets for sexual dysfunction in people

Female Rats are More Vulnerable to the Long-Term Consequences of Neonatal Inflammatory Injury

Premature infants are routinely exposed to invasive medical procedures during neonatal intensive care treatment that are largely performed in the absence of anesthetics or analgesics. Data collected to date suggest that exposure to early insult during this time of increased plasticity alters the development of the CNS and influences future pain responses. As previous studies examining the impact of neonatal injury on nociception have been conducted primarily in males, the potential adverse effects on females are not known. Therefore, the present studies were conducted to determine whether neonatal injury differentially impacts male and female sensory thresholds in adulthood. A short lasting inflammatory response was evoked in male and female rats on the day of birth with an injection of carrageenan (CGN; 1% or 2%) into the right hindpaw. Nociceptive thresholds were assessed using a noxious thermal stimulus at both adolescence (P40) and adulthood (P60). A more persistent inflammation was subsequently evoked in adult rats with an intraplantar injection of Complete Freund’s adjuvant (CFA). Neonatally injured females exhibited significantly greater hypoalgesia at P60, and displayed enhanced inflammatory hyperalgesia following re-injury in adulthood compared to neonatally injured males and controls. These results demonstrate that the long-term adverse effects of neonatal injury are exacerbated in females, and may contribute to the higher prevalence, severity and duration of pain syndromes noted in women compared to men

Nucleus Paragigantocellularis Afferents in Male and Female Rats: Organization, Gonadal Steroid Receptor Expression, and Activation During Sexual Behavior

The supraspinal regulation of genital reflexes is poorly understood. The brainstem nucleus paragigantocellularis (nPGi) of rats is a well-established source of tonic inhibition of genital reflexes. However, the organization, gonadal steroid receptor expression, and activity of nPGi afferents during sex have not been fully characterized in male and female rats. To delineate the anatomical and physiological organization of nPGi afferents, the retrograde tracer Fluoro-Gold (FG) was injected into the nPGi of sexually experienced male and female rats. Animals engaged in sexual behavior 1 hour before sacrifice. Cells containing FG, estrogen receptor-α (ERα), androgen receptor (AR), and the immediate-early gene product Fos were identified immunocytochemically. Retrograde labeling from the nPGi was prominent in the bed nucleus of the stria terminalis, paraventricular nucleus (PVN), posterior hypothalamus, precommissural nucleus, deep mesencephalic nucleus, and periaqueductal gray (PAG) of both sexes. Sex differences were observed in the caudal medial preoptic area (MPO), with significantly more FG+ cells observed in males, and in the PAG and inferior colliculus, where significantly more FG+ cells were observed in females. The majority of regions that contained FG+ cells also contained ERα or AR, indicating sensitivity to gonadal steroids. The proportions of FG+ cells that co-localized with sex-induced Fos was high in the PVN of both sexes and high in the MPO of males but low in the PAG of both sexes despite the large number of PAG-nPGi output neurons and Fos+ cells in both sexes. The characterization of these afferents will lead to a further understanding of the neural regulation of genital reflexes

Androgen and Estrogen (α) Receptor Localization on Periaqueductal Gray Neurons Projecting to the Rostral Ventromedial Medulla in the Male and Female Rat

The periaqueductal gray (PAG) is involved in many gonadal steroid-sensitive behaviors, including responsiveness to pain. The PAG projects to the rostral ventromedial medulla (RVM), comprising the primary circuit driving pain inhibition. Morphine administered systemically or directly into the PAG produces greater analgesia in male compared to female rats, while manipulation of gonadal hormones alters morphine potency in both sexes. It is unknown if these alterations are due to steroidal actions on PAG neurons projecting to the RVM. The expression of androgen (AR) and estrogen (ERα) receptors in the PAG of female rats and within this descending inhibitory pathway in both sexes is unknown. The present study used immunohistochemical techniques (1) to map the distribution of AR and ERα across the rostrocaudal axis of the PAG; and (2) to determine whether AR and/or ERα were colocalized on PAG neurons projecting to the RVM in male and female rats. AR and ERα immunoreactive neurons (AR-IR, ERα-IR) were densely distributed within the caudal PAG of male rats, with the majority localized in the lateral/ventrolateral PAG. Females had significantly fewer AR-IR neurons, while the quantity of ERα was comparable between thesexes. In both sexes, approximately 25-50% of AR-IR neurons and 20-50% of ERα-IR neurons were retrogradely labeled. This study provides direct evidence of the expression of steroid receptors in the PAG and the descending pathway driving pain inhibition in both male and female rats and may provide a mechanism whereby gonadal steroids modulate pain and morphine potency