Judith L. Steinman

Spinal cord monoamines modulate the antinociceptive effects of vaginal stimulation in rats

Abstract Perispinal administration (into the lumbar intrathecal space) of phentolamine (40 &mgr;g), an &agr;‐adrenergic receptor blocking agent, reduced the analgesic effect of vaginal stimulation by 39.7% (measured by tail flick latency) and 57.1% (measured by vocalization threshold) as compared to controls. Perispinal administration of methysergide (10 &mgr;g), a serotoninergic receptor blocking agent, reduced the analgesic effect of vaginal stimulation by 48.5% (measured by vocalization threshold), although it did not significantly affect the tail flick measure. In a separate experiment, vaginal stimulation activated the release of norepinephrine and serotonin into a superfusate of the spinal cord. During vaginal stimulation, levels of norepinephrine and serotonin increased about 2‐fold above resting levels. These findings indicate that vaginal stimulation releases norepinephrine and serotonin into the spinal cord, thereby exerting an analgesic effect.

Differential roles of hypogastric and pelvic nerves in the analgesic and motoric effects of vaginocervical stimulation in rats

Bilateral transection of the pelvic and/or hypogastric nerves, which convey afferent activity from the reproductive tract, was performed to ascertain the role of these nerves in the analgesic and motoric effects of vaginocervical mechanostimulation (VS) in rats. Two indices of analgesia were used: tail flick latency to radiant heat (TFL) and vocalization threshold to electrical shock of the tail (Voc-T). Nerve cuts were performed at least one week prior to behavioral testing. Bilateral transection of both the pelvic and hypogastric nerves eliminated the analgesic effects of VS on the TFL and Voc-T tests. Bilateral transection of only the pelvic nerves reduced the number of rats showing maximal VS-induced elevation in TFL, without altering the effect of VS on Voc-T. By contrast, bilateral transection of only the hypogastric nerves attenuated the Voc-T-elevating effect of VS, without reducing the effect of VS on elevating TFL. The effects of VS on producing immobility, hindlimb extension and blockage of hindlimb withdrawal to foot pinch were eliminated by combined bilateral pelvic and hypogastric neurectomy. However, bilateral transection of either nerve alone did not significantly alter the efficacy of VS in producing these effects. These findings indicate that the pelvic and hypogastric nerves contribute to the immobility- and extensor-inducing, and flexor-inhibiting effects of VS, and differentially mediate the analgesia-producing effects of VS.

The segmental distribution of afferent fibers from the vaginal cervix and hypogastric nerve in rats

Injections of horseradish peroxidase-wheat germ agglutinin (HRP-WGA) into the walls of the vagina and cervix (vaginocervical injections) of rats resulted in labeling of dorsal root ganglia (DRG) cells located at T11-L4 and L6-S2. In a second group of animals, exposure of the hypogastric nerve to HRP-WGA resulted in a similar bimodal distribution of labeled cells as compared to vaginocervical injections. In a third group, unilateral hypogastric nerve transection prior to injection of HRP-WGA into the vaginocervical walls resulted in a significant reduction in DRG cells labeled at T13, L1, L2, L6 and S1. Bilateral transection of the hypogastric nerves prior to vaginocervical injections eliminated labeled DRG cells at thoracolumbar levels but not at L6 and S1. Bilateral pelvic neurectomy reduced, but did not eliminate labeled DRG cells at L6 and S1 following vaginocervical injections. These results indicate that the hypogastric nerve constitutes a major sensory pathway from the vaginocervical walls to thoracic, lumbar and sacral levels of the spinal cord. The hypogastric nerve may subserve the transmission of noxious input from the vaginocervical walls as well as the activation of ascending spinal pathways involved in neuroendocrine reflexes during parturition.

Expression of c-fos protein in lumbosacral spinal cord in response to vaginocervical stimulation in rats.

The pattern of vaginocervical stimulation-evoked expression of the proto-oncogene c-fos in lumbar 5-sacral 1 segments of the spinal cord of ovariectomized adult rats was mapped using immunocytochemistry. A calibrated force of mechanostimulation was applied to the vaginal cervix of experimental animals and to the perineum of control animals while they were gently restrained. The number of cells expressing c-fos was significantly greater in the experimental than the control animals in laminae I, IV, V-VI and X. The implications of the present findings for elucidating the spinal pathways mediating the various behavioral, neuroendocrine and autonomic effects of vaginocervical stimulation (VS) are discussed.

Vaginocervical stimulation releases oxytocin within the spinal cord in rats.

Vaginocervical stimulation (VS) significantly elevated the concentration of oxytocin (OT) in spinal cord superfusates of 8 intact urethane-anesthetized rats measured 10–15 min after VS (median [interquartile range]: 1.7 [1.00–3.37] pg/ml) compared to that measured 10–15 min before VS (1.1 [1.01–1.40] pg/ml). When VS was administered once (n = 8), it produced a 55% increase over baseline values; when administered a second time 45 min later (n = 6), it produced only a 22% increase over pre-VS values. The effects of estrogen on the VS-induced release of OT were then investigated using ovariectomized rats that were treated either with estradiol benzoate (EB; 10 µg/100 g bw) (n = 6) or with an oil vehicle (n = 6) subcutaneously for 3 days. The EB treatment significantly elevated the basal levels of OT released into spinal cord superfusates above vehicle control levels. Within 5–10 min after the onset of VS, OT concentrations in the superfusates were significantly higher in EB-treated than in vehicle-treated rats. The vehicle-treated rats did not show a significant elevation in OT concentration following VS. To rule out the possibility that the posterior pituitary gland was the source of this OT, the effect of hypophysectomy (HYPOX) was assessed on the VS-induced release of OT into spinal cord superfusates and plasma. The concentration of OT in spinal cord superfusates of both the HYPOX (n = 5) and intact rats (n = 6) increased significantly from 5.8 [4.4–6.5] pg/ml pre-VS to 7.9 [6.7–10.3] pg/ml immediately after VS, and from 4.4 [3.8–5] pg/ml pre-VS to 5.1 [4.6–5.7] pg/ml immediately after VS, respectively. There was no significant difference in baseline levels of OT in cerebrospinal fluid between the two groups. By contrast, plasma OT levels, while significantly elevated in response to VS from 3.42 [2.9–5.34] pg/ml baseline to 7.25 [5.33–15.77] pg/ml in the intact group, failed to respond significantly to VS in the HYPOX group (n = 5). The present findings provide evidence of a direct estrogen-dependent release of OT within the spinal cord in response to VS, presumably via descending oxytocinergic neurons.

Genital Stimulation as a Trigger for Neuroendocrine and Behavioral Control of Reproduction

Coital stimulation normally provokes ovulation in rabbits, cats, ferrets, mink, and voles,’ which are therefore termed “reflexive ovulators.” “Spontaneous ovulators” (e.g., rats, humans, mice, cattle) normally ovuiate in response to cyclical changes in hormone levels: but under certain conditions, such as estrogen-induced mating in die~trus,~ barbiturate suppression of the neural activity preceding spontaneous ovulation: or persistent estrus produced by constant light: rats may ovulate reflexively. Thus the difference between reflexively and spontaneously ovulating species appears to be one of degree.6 In rabbits, coital stimulation depletes luteinizing hormone (LH) from the anterior pituitary within 15 min of mating’ and in rats made to be in persistent estrus by exposure to continuous light, coital stimulation increases plasma LH within 10 min.5 Artificial vaginal stimulation elevates plasma LH within 30 min in anesthetized rats?

Pelvic neurectomy blocks oxytocin-facilitated sexual receptivity in rats

The effect of bilateral pelvic, hypogastric, or pudendal neurectomy on oxytocin-induced facilitation of lordosis behavior was examined in ovariectomized, estrogen, and progesterone primed female rats. Oxytocin-induced facilitation of lordosis behavior was significantly decreased following bilateral pelvic, or combined bilateral pelvic and hypogastric neurectomy. Hypogastric only or pudendal only neurectomy had no effect on the facilitation of lordosis behavior after systemic administration of oxytocin. These results suggest that the integrity of the pelvic nerve is necessary and sufficient to mediate the stimulatory effects of systemically administered oxytocin on receptivity.

Antagonism of morphine analgesia by nonopioid cold-water swim analgesia: Direct evidence for collateral inhibition

The demonstrated existence of opioid and nonopioid forms of pain control has raised questions as to how they interact. Previous indirect evidence suggests that activation of one system inhibited the activation of the other. The present study assessed this directly using morphine as an opiate form of analgesia and continuous cold-water swims (CCWS, 4 degrees C, 2 min) as the nonopioid form. A significant reduction in morphine (8 mg/kg, SC) analgesia on the tail-flick test was observed if rats were acutely exposed to CCWS immediately prior to morphine administration. The inability of naloxone (10 mg/kg, SC) to reduce CCWS analgesia verified its nonopioid nature. The antagonism of morphine (3 mg/kg, SC) analgesia was greater following preexposure to 2 min of CCWS than 1 min of CCWS. CCWS was also more effective in antagonizing analgesia induced by the 3 mg/kg than the 8 mg/kg dose of morphine. The antagonism of morphine analgesia by CCWS was dependent upon the temporal patterning of stimulus presentation: exposure to CCWS 20 or 60 min prior to morphine failed to alter subsequent morphine analgesia. A significant reduction in analgesia induced by intraperitoneal administration of morphine (10 mg/kg) was also observed when CCWS was presented immediately prior to injection, suggesting that pharmacokinetic factors such as altered drug absorbance by CCWS-induced vasoconstriction do not appear to explain these effects. These data provide direct support for the existence of collateral inhibitory mechanisms activated by CCWS and morphine, and suggests that these opioid and nonopioid forms of analgesia do not function synergistically, but instead involve some form of hierarchical order.

Forebrain and hindbrain involvement of neuropeptide Y in ingestive behaviors of rats

Neuropeptide Y (NPY) is an extremely potent orexigenic agent. These studies demonstrate that the effect of NPY on food and water intake are seen after infusion into either the third (3V) or fourth (4V) ventricle and that this is a specific effect, as it was not seen with the deaminated form. There was a nonsignificant tendency for lateral midbrain knife cuts to food intake. Both 3V and 4V NPY infusions showed an attenuated increases in food intake at 1 but not 2 h following NPY infusion in the lateral knife cut rats compared to the sham controls. Medical knife cuts resulted in significantly greater food intake in the basal state and a lesser increase in food intake in response to NPY infused into the 3V. These midbrain data suggest a role for both medical and lateral fibers in mediating the effects of NPY on food intake. Lateral fibers appear to be more important, but their transection only delays the time of onset of the stimulating effect of NPY to the second hour. Lateral knife cuts virtually abolish the effect of 4V NPY on stimulating water intake. 3V NPY in the presence of NPY has a less clear effect at 1 h, but mildly attenuated the NPY effect on water intake at 2 h in lateral knife cut rats. Medial knife cuts slightly attenuate the effect of 3V NPY on water intake. However, medial knife cuts markedly increased basal water ingestion. These studies demonstrate the importance of neuronal communications between third and fourth ventricle associated structures in the modulation of ingestive behavior.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential effects ofp-chlorophenylalanine on indoleamines in brainstem nuclei and spinal cord of rats. I. Biochemical and behavioral analysis

The involvement of endogenous serotonergic pathways in the mediation of antinociception has been indicated by electrophysiological, pharmacological and behavioral experiments. However, manipulation of the indole pathway, either by lesioning of raphe nuclei or drug intervention, often produces disparate results. In particular, serotonin (5-HT) synthesis inhibition with p-chlorophenylalanine (PCPA) has been reported to produce either hyperalgesia or analgesia, depending upon the type of pain measurement examined. In the present study, we sought to evaluate the effects of PCPA on (1) behavioral responses to noxious stimulation, and (2) levels of serotonin, tryptophan and 5-hydroxyindoleacetic acid (5-HIAA) in raphe nuclei (pallidus, obscurus, magnus and dorsalis) and spinal cord regions by HPLC with electrochemical detection. Treatment of rats with 400 or 600 mg/kg of PCPA for 3 consecutive days resulted in significant elevations in pain thresholds assessed by tail withdrawal from radiant heat as well as vocalization to electric shock of the tail. The effect of PCPA on vocalization threshold was particularly striking, for the majority of animals showed a nociceptive-specific attenuation of this response. Although the PCPA induced changes in indole content of the various raphe nuclei were not unequivocally dose-dependent, differential reductions of serotonin and 5-HIAA were clearly detected in the various raphe regions. Nuclei raphe pallidus and obscurus were depleted of 5-HT and 5-HIAA to the greatest extent, whereas levels detected in nuclei raphe magnus and dorsalis were reduced by 30-40% from control values. Metabolism of 5-HT and 5-HIAA appeared unaffected by PCPA in all regions examined except the dorsal portion of the spinal cord. These findings collectively suggest that the effects of PCPA are not uniform throughout the central nervous system and raise the possibility that discrepancies in the behavior literature may be attributed to drug-induced changes in some, but not all serotonergic pathways.