Effects of space flight on the immunohistochemical demonstration of connexin 26 and connexin 43 in the postpartum uterus of rats.

The effect of space flight in a National Aeronautics and Space Administration shuttle was studied in pregnant rats. Rats were launched on day 11 of gestation and recovered on day 20 of gestation. Pregnancy was allowed to proceed to term and rats delivered vaginally on days 22-23, although flight animals required more labour contractions to complete the delivery process. Pups were placed with foster dams and connexin 26 and 43 were examined in the uterus of flight animals approximately 3 h after delivery. Space flight did not affect uterine connexin 26, localized primarily in epithelial cells of the endometrium, but decreased connexin 43, the major gap junction protein in the myometrium. It is suggested that decreased connexin 43 alters synchronization and coordination of labour contractions, resulting in a requirement for more contractions to complete the delivery process.

The effects of space flight during gestation on rat uterine smooth muscle.

Pregnant rats were flown on a NASA space shuttle during days 11-20 of gestation and the effects of pregnancy maintenance and uterine myometrial smooth muscle were evaluated. There was no effect of space flight on fetal mass in rats examined at recovery on day 20 of gestation (G-20). Rats allowed to go to term delivered vaginally at days 22-23, but space flight significantly decreased pup mass at birth. Space flight did not alter myometrial smooth muscle volume at G-20 and postpartum when compared to flight delayed synchronous controls, but flight animals showed a 37% decrease in myometrial smooth muscle volume between G-20 and postpartum. In contrast, smooth muscle volume of the myometrium of controls, postpartum, was similar to G-20. We conclude that myometrial smooth muscle, which hypertrophies during space flight similar to controls, reacclimates acutely to earth's gravity between G-20 and parturition with a dramatic reduction in volume.

Effects of space flight on ovarian-hypophyseal function in postpartum rats.

The effect of space flight in a National Aeronautics and Space Administration (NASA) shuttle was studied in pregnant rats. Rats were launched on day 9 of gestation and recovered on day 20 of gestation. On day 20 of gestation, rats were unilaterally hysterectomized and subsequently allowed to go to term and deliver vaginally. There was no effect of space flight on pituitary and ovary mass postpartum. In addition, space flight did not alter healthy and atretic ovarian antral follicle populations, fetal wastage in utero, plasma concentrations of progesterone and luteinizing hormone (LH) or pituitary content of follicle stimulating hormone (FSH). Space flight significantly increased plasma concentrations of FSH and decreased pituitary content of LH at the postpartum sampling time. Collectively, these data show that space flight, initiated during the postimplantation period of pregnancy, and concluded before parturition, is compatible with maintenance of pregnancy and has minimal effects on postpartum hypophyseal parameters; however, none of the ovarian parameters examined was altered by space flight.

Effect of prior nicotine treatment on drug induced changes in serum LH concentrations in rats.

The effect of daily injections of nicotine on drug induced changes in LH secretion was investigated in male rats. Daily administration of nicotine for 7 days resulted in decreased basal serum LH concentrations. Nicotine treatment blocked naloxone induced LH release and reduced LHRH induced increases in serum LH. Clonidine induced increases in serum LH were not altered by nicotine treatment and haloperidol treatment did not alter nicotine induced decreases in serum LH. In an acute study nicotine blocked LH secretion induced by the long acting opioid antagonist naltrexone. Collectively these results indicate that opioidergic neurons are involved in the reduction in serum LH that occurs following nicotine. They also indicate that chronic nicotine treatment can reduce the pituitary gland response to LHRH.

Sampling of prenatal and postnatal offspring from individual rat dams enhances animal use without compromising development.

To assess prenatal and postnatal developmental status in the offspring of a group of animals, it is typical to examine fetuses from some of the dams as well as infants born to the remaining dams. Statistical limitations often arise, particularly when the animals are rare or especially precious, because all offspring of the dam represent only a single statistical observation; littermates are not independent observations (biologically or statistically). We describe a study in which pregnant laboratory rats were laparotomized on day 7 of gestation (GD7) to ascertain the number and distribution of uterine implantation sites and were subjected to a simulated experience on a 10-day space shuttle flight. After the simulated landing on GD18, rats were unilaterally hysterectomized, thus providing a sample of fetuses from 10 independent uteruses, followed by successful vaginal delivery on GD22, yielding postnatal samples from 10 uteruses. A broad profile of maternal and offspring morphologic and physiologic measures indicated that these novel sampling procedures did not compromise maternal well-being and maintained normal offspring development and function. Measures included maternal organ weights and hormone concentrations, offspring body size, growth, organ weights, sexual differentiation, and catecholamine concentrations.

NADPH-diaphorase-positive neurons innervating the rat ovary.

Nerve cell bodies projecting to the ovary were visualized in dorsal root ganglia (DRG) and paravertebral ganglia after application of the retrograde tracer Fluoro-gold to the superior ovarian and plexus nerves. The location of fluorescent cells in sections of ganglia was recorded and subsequently nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry was utilized to locate nitric oxide synthase (NOS) whose presence indicates sites where nitric oxide (NO) can be synthesized. No fluorescent nerve cell bodies in paravertebral ganglia were NADPH-diaphorase-positive. In contrast, numerous Fluoro-gold-labeled nerve cell bodies in DRG at segmental levels T12-L1 were NADPH-diaphorase positive. Thus, many sensory neurons projecting to the ovary contain NOS and presumably release NO. This gaseous messenger molecule may participate in modulation of ovarian function.

Dorsal rhizotomy does not alter rat estrous cycles or ovarian compensatory responses.

Rats were subjected to right side dorsal rhizotomy of spinal nerves T12-L2 and ipsi- or contralateral unilateral ovariectomy at estrus (day 1). Estrous cycles were recorded daily, and on day 15 the remaining ovary was removed and weighed. Antral follicles were counted in representative day 1 and day 15 ovaries. Dorsal rhizotomy did not affect estrous cycles during the period after surgery. Also dorsal rhizotomy did not alter ipsilateral ovarian hypertrophy or follicular activation following unilateral ovariectomy. Thus, although the major portion of the ovarian sensory innervation is via the lesioned segments, interruption of these segments centrally does not alter compensatory ovarian responses subsequent to unilateral ovariectomy.

Naltrexone treatment attenuates the inhibitory effect of nicotine treatment on serum LH in rats.

The hypothesis that endogenous opioids might have a role in mediating the suppressive effects of nicotine on serum LH concentrations in rats was investigated. Naltrexone treatment prevented the inhibitory effect of high doses of nicotine on serum LH concentrations. Nicotine treatment also prevented the stimulatory effect of naltrexone on serum LH concentrations. These results suggest that the inhibitory effects of nicotine on serum LH concentrations involve an opioidergic component.

Endocrine parameters associated with disruption of parturition after bilateral pelvic neurectomy.

Bilateral lesions of the pelvic nerve (BLPN) result in dystocia, but the processes which control this effect are not fully understood. Plasma progesterone, relaxin, and luteinizing hormone (LH) concentrations were measured in blood samples taken in the morning (AM) and evening (PM) of Days 20-23 of gestation from rats with BLPN or sham neurectomy. Ten of 11 sham-operated control animals delivered their entire litters by Day 23 of gestation, but animals with BLPN did not complete parturition by Day 23 when they were sacrificed. Progesterone concentrations were greater in rats with BLPN than in sham-operated rats on Day 20 PM and Day 21 AM, but hormone concentrations declined to minimal values by Day 22 in both groups. Relaxin concentrations were greater in rats with BLPN than in sham-operated rats on Day 21 PM. Thereafter, relaxin concentrations decreased to reach minimum values on Day 23 in both groups. LH concentrations were low throughout the period of study in rats with BLPN; however, a postpartum LH surge was detected in all sham-operated animals. Data from this study indicate that the pelvic nerve does not control parturition by modulating serum relaxin and progesterone concentrations; however, these data suggest that impulses carried by the pelvic nerve influence ovarian secretion of these hormones. In addition, these data indicate that the pelvic nerve transmits stimuli from the cervix to the hypothalamus to facilitate the postpartum LH surge.

Effects of peripheral nerve lesions during pregnancy on parturition in rats.

Bilateral section of either the sensory or motor branch of the pelvic nerve or pudendal nerve was performed in rats on days 8-10 of pregnancy, and the effects on delivery were observed. Bilateral resection of the sensory branch of the pelvic nerve reduced the number of live pups per litter, and increased the number of stillbirths and the number of fetuses retained in utero per litter at day 24. Sectioning motor components of the pelvic nerve, or both motor and sensory components of the pudendal nerve, had no effects on delivery in rats. We conclude that of the peripheral nerves evaluated in this study, only the sensory branch of the pelvic nerve is required for normal vaginal delivery in this species.

Direct electrical stimulation of the superior ovarian nerve in rats causes an increase in neuronal activity in the ipsilateral ovarian plexus nerve.

Ovarian blood flow and neuronal activity in the ovarian plexus nerve (OPN) were monitored before, during and after direct electrical stimulation of the central cut end of the ipsilateral superior ovarian nerve (SON). Blood flow did not change during the observation period. Stimulation of the SON increased the frequency of action potentials in the OPN, suggesting an ovarian reflex pathway mediated by the SON and the OPN.

Afferent fibers in the reproductive system and pelvic viscera of female rats: anterograde tracing and immunocytochemical studies.

Innervation of the female reproductive system provides an important signal for a variety of neuroendocrine reflexes and behaviors in the female rat. Although some studies suggest that afferent feedback from the gonads is involved in the hypothalamic control of gonadal function and pituitary hormone release, the extent and function of afferent feedback from the gonads in these neuroendocrine reflexes has yet to be clarified. Deafferentation studies have provided only partial support for the afferent control of the gonads. Some studies even suggest functional asymmetries in the neural control of the gonads, but knowledge regarding the neuroanatomical substrate for these possible neurogonadal interactions remains incomplete. Studies with retrograde tract tracers indicate that the ovaries receive a substantial afferent supply from lower thoracic-upper lumbar dorsal root ganglia. Despite stringent precautions to prevent diffusion of tracers following injections into the ovary or related nerves, many of the retrogradely labeled cell bodies identified by these studies may represent an overestimation of the extent of afferent innervation. We have reexamined the afferent innervation of the female reproductive tract by means of the anterograde transport of horseradish peroxidase (HRP) from thoracic, lumbar and sacral dorsal root ganglion to pelvic visceral organs and have studied the effects of unilateral ganglionectomy on substance P containing fibers in the ovary, oviduct and uterus. The neuroanatomical results show that the T13 and L1 dorsal root ganglia provide major afferent innervation to the cranial portion of the reproductive tract and the L6 and S1 dorsal root ganglia provide primary afferent fibers to the caudal portion of the reproductive tract as well as the bladder, rectum and perineum.(ABSTRACT TRUNCATED AT 250 WORDS)

Substance P- and vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers in relation to ovarian postganglionic perikarya in para- and prevertebral ganglia: evidence from combined retrograde tracing and immunocytochemistry.

Retrograde neuronal tracing with the fluorescent dye True Blue and immunocytochemistry were utilized to examine postganglionic sympathetic neurons in para- and prevertebral ganglia projecting to the rat ovary. Perikarya in both ganglia were labeled with True Blue after application of the tracer to either the superior ovarian or ovarian plexus nerve. After application of True Blue to the superior ovarian nerve, 17% of the labeled cells in paravertebral ganglia were immunoreactive for vasoactive intestinal polypeptide. In contrast, after application of True Blue to the ovarian plexus nerve, approximately 1% of the labeled cells in paravertebral ganglia were immunoreactive for the same polypeptide. Some vasoactive intestinal polypeptide-immunoreactive perikarya in paravertebral ganglia were not labeled with True Blue. In some cases, substance P- and vasoactive intestinal polypeptide-immunoreactive fibers were closely apposed to True Blue-labeled perikarya in para- and prevertebral ganglia. Paravertebral vasoactive intestinal polypeptide-immunoreactive perikarya projecting to the ovary presumably participate directly in the control of various ovarian functions. Substance P- and vasoactive intestinal polypeptide-immunoreactive fibers closely apposed to perikarya projecting to the ovary may participate indirectly in the control of various ovarian functions by affecting the activity of ovarian postganglionic neurons.

Anatomical localization of afferent and postganglionic sympathetic neurons innervating the rat ovary.

The fluorescent retrograde tracer, True blue, was applied directly to either the superior ovarian nerve (SON) or the ovarian plexus nerve (OPN) in the rat. Afferent perikarya were located in lower thoracic-upper lumbar dorsal root ganglia and projected to the ovary via both nerve routes. Postganglionic sympathetic efferent perikarya were located in both prevertebral and thoracolumbar paravertebral ganglia and also utilized both the SON and OPN to reach the ovary. The significance of the dual origin of postganglionic sympathetic neurons innervating the rat ovary is not known.

Peripheral pathways for neuropeptide Y- and cholecystokinin-8-immunoreactive nerves innervating the rat ovary.

The superior ovarian nerve (SON) or plexus nerve (PN) innervating the rat ovary was transected separately or in combination and the effects of these nerve lesions on intra-ovarian NPY- or CCK-8-immunoreactive nerve fibers was evaluated. Transection of the SON did not affect NPY or CCK-8-immunoreactive nerve fibers. In contrast, section of the PN eliminated nerve fibers immunoreactive for these neuropeptides. This study demonstrates that both NPY- and CCK-8-immunoreactive nerve fibers reach the ovary via the PN.

Neuropeptides in sensory perikarya projecting to the rat ovary.

Afferent perikarya in dorsal root ganglia (DRG) at the T13 and L1 segmental levels projecting to the rat ovary were identified by utilizing the fluorescent retrograde tracer true blue (TB). Subsequently, TB-labeled ovarian afferent perikarya in DRG were examined for vasoactive intestinal polypeptide (VIP), substance P (SP), cholecystokinin-8 (CCK-8), neuropeptide Y (NPY), and somatostatin (SOM) immunoreactivity and for the presence of fluoride-resistant acid phosphatase (FRAP) enzyme activity. Of the ovarian afferent perikarya at the T13 and L1 segmental levels, 20.5% displayed VIP immunoreactivity, 23.8% displayed SP immunoreactivity, and 43.1% were immunoreactive for CCK-8. No ovarian afferent perikarya contained SOM or NYP immunoreactivity or FRAP activity. It is suggested that different neuropeptides may participate in modulation of specific ovarian functions.

Neuropeptide Y and somatostatin immunoreactive perikarya in preaortic ganglia projecting to the rat ovary.

Postganglionic perikarya in preaortic ganglia projecting to the ovary of the rat were identified utilizing the retrograde fluorescent tracer, true blue. Ovarian perikarya were subsequently examined for neuropeptide Y and somatostatin immunoreactivity. True blue-labelled ovarian postganglionic perikarya were distributed in the coeliac ganglion and in smaller ganglia located at the origins of the renal and ovarian arteries. In the coeliac ganglia, 74.4 +/- 18.8% of true blue-labelled ovarian perikarya were immunoreactive to neuropeptide Y while 37.4 +/- 9.6% were immunoreactive to somatostatin. In the inferior smaller ganglia, 72.2 +/- 18.7% and 2.2 +/- 2.2% of the true blue-labelled ovarian perikarya were immunoreactive to neuropeptide Y and somatostatin respectively. It is suggested that neuropeptide Y and somatostatin participate in the modulation of ovarian function.

Utilization of a polyethylene enclosure for nerve tracing studies in the rat ovary.

A commonly used technique for mapping neuronal connections in the peripheral nervous system involves the injection of a tracer substance into a peripheral organ. The tracer is taken up by nerve terminals at the injection site and transported retrogradely to motor or sensory perikarya where it can be visualized. One of the pitfalls of this technique is that tracer may leak from the injection site, enter nerve terminals adjacent to the injection area and thus cause erroneous interpretations. Similarly, some tracers may enter blood vessels and be taken up by nerve cell bodies indiscriminately. In order to circumvent these potential problems in studies on the innervation of the rat ovary, a polyethylene enclosure for the organ was fabricated from a BEEM capsule. The retrograde tracer, True blue, was then applied topically on the ovarian surface inside the enclosure. In chronic experiments, specifically labeled ovarian nerve cell bodies were identified in dorsal root and preaortic ganglia. Utilization of this simple and effective enclosure eliminates the false-positive and non-specific labeling of perikarya due to diffusion and blood-borne transport of the tracer from the injection area.

Convergence of sensory processes from the heart and left ulnar nerve onto a single afferent perikaryon: a neuroanatomical study in the rat employing fluorescent tracers.

After injection of true blue (TB) into the pericardial sac and nuclear yellow (NY) into the left ulnar nerve of rats, most perikarya in the left eighth cervical and first thoracic dorsal root ganglia are single-labeled with either TB or NY. However, 7.1-14.7% of the sensory perikarya from the cardiac area labeled with TB are simultaneously labeled with NY. This finding indicates that some primary sensory perikarya receive processes from both the left arm and the heart. In addition to other mechanisms that may be operative in the referral of pain of cardiac origin to the medial left arm, these results suggest that sensory neurons with dichotomizing somatic and visceral peripheral processes may also contribute to this phenomenon.

Effect of abdominal vagotomy at proestrus on ovarian weight, ovarian antral follicles, and serum levels of gonadotropins, estradiol, and testosterone in the rat.

The effects of abdominal vagotomy at proestrus on ovarian weight and antral follicles greater than 150 microns diameter and on serum levels of gonadotropins and testosterone were assessed 24 and 48 h and 4 and 8 days after surgery. Serum levels of estradiol were assessed at 4 and 8 days. Vagotomy increased ovarian weight at 48 h, decreased ovarian weight at 4 days, but had no effect by day 8. Vagotomy increased healthy antral follicles 151-394 microns diameter at 24 and 48 h and increased atresia in this size range at 4 and 8 days. Vagotomy decreased healthy follicles 151-384 microns at day 8. Vagotomy decreased healthy follicles 395-570 microns at 24 h and decreased atretic follicles at 48 h. Vagotomy decreased the largest (over 570 micron diameter) healthy follicles at 24 h and 8 days. Vagotomy decreased basal serum LH levels at 48 h and 8 days. (In contrast, vagotomy increased FSH at 24 h). There was no effect on blood levels of estradiol and testosterone. These findings are discussed in relation to the hypothesis that the vagus nerve is a component of the hypothalamo-hypophyseal-ovarian axis.