Immunocytochemical distribution of catecholamine-synthesizing neurons in the hypothalamus and pituitary gland of pigs: tyrosine hydroxylase and dopamine-beta-hydroxylase.

This study describes the distribution of catecholaminergic neurons in the hypothalamus and the pituitary gland of the domestic pig, Sus scrofa, an animal that is widely used as an experimental model of human physiology in addition to its worldwide agricultural importance. Hypothalamic catecholamine neurons were identified by immunocytochemical staining for the presence of the catecholamine synthesizing enzymes, tyrosine hydroxylase and dopamine-beta-hydroxylase. Tyrosine hydroxylase-immunoreactive perikarya were observed in the periventricular region throughout the extent of the third ventricle, the anterior and retrochiasmatic divisions of the supraoptic nucleus, the suprachiasmatic nucleus, the ventral and dorsolateral regions of the paraventricular nucleus and adjacent dorsal hypothalamus, the ventrolateral arcuate nucleus, and the posterior hypothalamus. Perikarya ranged from parvicellular (10-15 microns) to magnocellular (25-50 microns) and were of multiple shapes (rounded, fusiform, triangular, or multipolar) and generally had two to five processes with branched arborization. No dopamine-beta-hydroxylase immunoreactive perikarya were observed within the hypothalamus or in the adjacent basal forebrain structures. Both tyrosine hydroxylase- and dopamine-beta-hydroxylase-immunoreactive fibers and punctate varicosities were observed throughout areas containing tyrosine hydroxylase perikarya, but dopamine-beta-hydroxylase immunoreactivity was very sparse within the median eminence. Within the pituitary gland, only tyrosine hydroxylase fibers, and not dopamine-beta-hydroxylase immunoreactive fibers, were located throughout the neurohypophyseal tract and within the posterior pituitary in both pars intermedia and pars nervosa regions. Generally, the location and patterns of both catecholamine-synthesizing enzymes were similar to those reported for other mammalian species except for the absence of the A15 dorsal group and the very sparse dopamine-beta-hydroxylase immunoreactive fibers and varicosities in the median eminence in the pig. These findings provide an initial framework for elucidating behavioral and neuroendocrine species differences with regard to catecholamine neurotransmitters.

Immunocytochemical localization of the catecholamine-synthesizing enzymes, tyrosine hydroxylase and dopamine-beta-hydroxylase, in the hypothalamus of cattle.

Immunocytochemical staining for the presence of catecholamine synthesizing enzymes, tyrosine hydroxylase and dopamine beta-hydroxylase, was used to characterize the regional distribution of catecholaminergic neurons in the hypothalamus and adjacent areas of domestic cattle, Bos taurus. In steers, heifers and cows, tyrosine hydroxylase-immunoreactive perikarya was located throughout periventricular regions of the third cerebral ventricle, in both anterior and retrochiasmatic divisions of the supraoptic nucleus, suprachiasmatic nucleus, and ventral and dorsolateral regions of the paraventricular nucleus, dorsal hypothalamus, ventrolateral aspects of the arcuate nucleus, along the ventral hypothalamic surface between the median eminence and optic tract, and in the posterior hypothalamus. Immunostained perikarya ranged from small (10-20 microns, parvicellular) to large (30-50 microns, magnocellular) and were of multiple shapes: round, triangular, fusiform or multipolar, often with 2-5 processes of branched arborization. There were no dopamine-beta-hydroxylase immunoreactive perikarya observed within the hypothalamus and adjacent structures. However, both tyrosine hydroxylase and dopamine-beta-hydroxylase immunoreactive fibers and punctate varicosities were observed throughout regions of tyrosine hydroxylase immunoreactivity perikarya. Generally, the location and pattern of hypothalamic tyrosine hydroxylase immunoreactivity and dopamine-beta-hydroxylase immunoreactive were similar to those reported for most other large brain mammalian species, however, there were several differences with commonly used small laboratory animals. These included intense tyrosine hydroxylase immunoreactivity of perikarya within the retrochiasmatic division of the supraoptic nucleus (ventral A15 region), the absence of tyrosine hydroxylase immunoreactive perikarya below the anterior commissure or within the bed nucleus of stria terminalis (absence of the dorsal A15 region), an abundance of tyrosine hydroxylase immunoreactive perikarya within the ependymal layer of the median eminence, heavy innervation of the arcuate nucleus with dopamine-beta-hydroxylase immunoreactive fibers and varicosities, and the paucity of dopamine-beta-hydroxylase immunoreactive throughout the median eminence.

Plasma cortisol and beta-endorphin in horses subjected to electro-acupuncture for cutaneous analgesia.

Electro-acupuncture (EA) treatment of horses to induce cutaneous analgesia also increased plasma concentrations of beta-endorphin (beta-EP) and cortisol. The magnitude of these increases did not relate consistently to the degree of EA-induced analgesia. Respiration and heart rates were also markedly increased during EA treatment. Intact female horses had higher packed cell volume and plasma beta-EP as well as lower plasma total protein than castrated male horses. Plasma cortisol, heart rate, and respiration rate did not differ significantly between sexes. None of the parameters measured before or during EA treatment provided an explanation for the differential cutaneous analgesia which depended on sex of subject and locus of stimulation as reported elsewhere.

Wound healing and angiogenic properties of supernatants from Lactobacillus cultures.

Extracts or supernatants from cultures of Lactobacilli are used for their medicinal effects, including wound healing and immune system stimulating activity. We have studied the in vivo and in vitro effects of supernatants from bacterial cultures of two strains of Lactobacillus (LS) on tissue repair and angiogenesis. Subcutaneous injection of LS into rodent ears led to proliferation of blood vessels that also exhibited strong immunostaining for Flk-1 receptor. Some inflammatory cells were scattered among the blood vessels. The continuous influx of polymorphonuclear leukocytes (PMNs) and macrophages into transcutaneous wounds in mice treated with LS resulted in prolonged inflammatory phase of wound healing and delayed wound closure, including reepithelialization. Subcutaneous injection of Matrigel impregnated with LS into the abdominal wall led to rapid and transient influx of PMNs in the vicinity of the gel. LS stimulated the proliferation of murine macrophage J774.A1 cell line and porcine lymphocytes but not that of murine fibroblast AKR-2B cells. LS also induced production of TNF-alpha by J774.A1 cells and by porcine kidney epithelial LLC-PK1 cells. LS did not appear to have an effect on collagen production. In conclusion, our study demonstrates the potential of LS to function as a stimulator of the inflammatory stage of tissue repair, TNF-alpha production, and of angiogenesis.

Catecholaminergic region A15 in the bovine and porcine hypothalamus.

Magnocellular perikarya within the retrochiasmatic division of the supraoptic nucleus of bovine and porcine hypothalami were immunoreactive (ir) with antiserum against tyrosine hydroxylase (TH), but not dopamine-beta-hydroxylase (DBH). Few cells in this region were also immunoreactive for vasopressin (VP) or oxytocin (OT). In contrast, the main division of the supraoptic nucleus contained numerous perikarya immunoreactive for VP and OT, but not TH nor DBH. Both the retrochiasmatic and principal divisions of the supraoptic nuclei contained TH- and DBH-ir fibers and varicosities. This region in bovine and porcine hypothalami corresponds to the ventral A15 catecholaminergic (dopamine-producing) cell group.

Central nervous system peptide and amino acid modulation of luteinizing hormone and prolactin secretion in the pig.

We recently demonstrated that pulsatile LH secretion is associated with pulsatile gonadotropin releasing hormone (GnRH) in the pig. Endogenous opioid peptide (EOP) inhibition of pulsatile LH and prolactin (PRL) secretion is dependent on reproductive status and development of this EOP system is a brain maturational process independent of the ovary. Once sexual maturation has occurred, EOP then become part of a progesterone dependent system and EOP inhibit a noradrenergic component of this system. During lactation, EOP also inhibit pulsatile LH, but stimulate PRL secretion. N-methyl-d,l-aspartate (NMA), an agonist of the excitatory amino acids (EAA), aspartate and glutamate, suppressed LH secretion in gilts pretreated with progesterone or vehicle. Both the EOP agonist, morphine (MOR), and the EOP antagonist, naloxone (NAL), delayed emergence and time to maximum serum LH concentration of the estradiol-induced LH surge in prepuberal and mature gilts, respectively. Therefore, EOP may normally have both a permissive as well as an inhibitory role in the LH surge mechanism. Although a norepinephrine synthesis inhibitor failed to alter basal PRL secretion, the PRL increase after NAL was suppressed in progesterone-treated ovariectomized (OVX) gilts. NAL suppressed the PRL response to NMA in OVX gilts pretreated with oil vehicle or progesterone, indicating that NMA stimulation of PRL secretion is mediated through the EOP system.

Opioid modulation of gonadotropin releasing hormone release from the hypothalamic preoptic area in the pig.

Two experiments (Exp) were conducted to examine in vitro the release of gonadotropin releasing hormone (GnRH) from the hypothalamus after treatment with naloxone (NAL) or morphine (MOR). In Exp 1, hypothalamic-preoptic area (HYP-POA) collected from 3 market weight gilts at sacrifice and sagittally halved were perifused for 90 min prior to a 10 min pulse of morphine (MOR; 4.5 x 10(-6) M) followed by NAL (3.1 x 10(-5) M) during the last 5 min of MOR (MOR + NAL; n = 3). The other half of the explants (n = 3) were exposed to NAL for 5 min. Fragments were exposed to KCl (60 mM) at 175 min to assess residual GnRH releasability. In Exp 2, nine gilts were ovariectomized and received either oil vehicle im (V; n = 3); 10 micrograms estradiol-17 beta/kg BW in 42 hr before sacrifice (E; n = 3); .85 mg progesterone/kg BW in twice daily for 6 d prior to sacrifice (P4; n = 3). Blood was collected to assess pituitary sensitivity to GnRH (.2 microgram/kg BW) on the day prior to sacrifice. On the day of sacrifice HYP-POA explants were collected and treated as described in Exp 1 except tissue received only NAL. In Exp 1, NAL increased (P < .05) GnRH release. This response to NAL was attenuated (P < .05) by coadministration of MOR. Cumulative GnRH release after NAL was greater (P < .05) than after MOR + NAL. All tissues responded similarly to KCl with an increase (P < .05) in GnRH release.(ABSTRACT TRUNCATED AT 250 WORDS)

Associated luteinizing hormone-releasing hormone and luteinizing hormone secretion in ovariectomized gilts.

The secretion of luteinizing hormone-releasing hormone (LHRH) and its temporal association with pulses of luteinizing hormone (LH) was examined in ovariectomized prepuberal gilts. Push-pull cannulae (PPC) were implanted within the anterior pituitary gland and LHRH was quantified from 10 min (200 microliters) perfusate samples. Serum LH concentrations were determined from jugular vein blood obtained at the midpoint of perfusate collection. Initial studies without collection of blood samples, indicated that LHRH secretion in the ovariectomized gilt was pulsatile with pulses comprised of one to three samples. However, most pulses were probably of rapid onset and short duration, since they comprised only one sample. Greater LHRH pulse amplitudes were associated with PPC locations within medial regions of the anterior pituitary close to the median eminence. In studies which involved blood collection, LH secretion was not affected by push-pull perfusion of the anterior pituitary gland in most gilts, however, adaptation of pigs to the sampling procedures was essential for prolonged sampling. There was a close temporal relationship between perfusate LHRH pulses and serum LH pulses with LHRH pulses occurring coincident or one sample preceding serum LH pulses. There were occasional LHRH pulses without LH pulses and LH pulses without detectable LHRH pulses. These results provide direct evidence that pulsatile LHRH secretion is associated with pulsatile LH secretion in ovariectomized gilts. In addition, PPC perfusion of the anterior pituitary is a viable procedure for assessing hypothalamic hypophyseal neurohormone relationships.

Effects of suckling and of a long interval after ovariectomy on hypothalamo-hypophyseal responsiveness to naloxone, morphine and GnRH in beef cows.

The response of serum luteinizing hormone (LH) to morphine, naloxone and gonadotropin-releasing hormone (GnRH) in ovariectomized, suckled (n=4) and nonsuckled (n=3) cows was investigated. Six months after ovariectomy and calf removal, the cows were challenged with 1mg, i.v. naloxone/kg body weight and 1 mg i.v. morphine/kg body weight in a crossover design; blood was collected at 15-minute intervals for 7 hours over a 3-day period. To evaluate LH secretion and pituitary responsiveness, 5 microg of GnRH were administered at Hour 6 on Day 1. On Days 2 and 3, naloxone or morphine was administered at Hour 3, followed by GnRH (5 microg/animal) at Hour 6. Mean preinjection LH concentrations (3.6 +/- 0.2 and 4.7 +/- 0.2 ng/ml), LH pulse frequency (0.6 +/- 0.1 and 0.8 +/- 0.1 pulses/hour) and LH pulse amplitude (2.9 +/- 0.5 and 2.9 +/- 0.6 ng/ml) were similar for suckled and nonsuckled cows, respectively. Morphine decreased (P < 0.01) mean serum LH concentrations (pretreatment 4.2 +/- 0.2 vs post-treatment 2.2 +/- 0.2 ng/ml) in both suckled and nonsuckled cows; however, mean serum LH concentrations remained unchanged after naloxone. Nonsuckled cows had a greater (P < 0.001) LH response to GnRH than did suckled cows (area of response curve: 1004 +/- 92 vs 434 +/- 75 arbitrary units). We suggest that opioid receptors are functionally linked to the GnRH secretory system in suckled and nonsuckled cows that had been ovariectomized for a long period of time. However, gonadotropin secretion appears not to be regulated by opioid mechanisms, and suckling inhibits pituitary responsiveness to GnRH in this model.

Effects of endophyte-infected fescue on concentrations of prolactin in blood sera and the anterior pituitary and concentrations of dopamine and dopamine metabolites in brains of steers.

An experiment was conducted to determine if the decrease in circulating concentrations of prolactin in cattle consuming endophyte (Acremonium coenophialum) -infected tall fescue (Festuca arundinacea) was associated with changes in prolactin concentrations in the anterior pituitary and concentrations of dopamine (DA) and its metabolites 3,4-dihydroxyphenyl-acetic acid (DOPAC) and homovanillic acid (HVA) in the stalk median eminence (SME), preoptic area (POA) and hypothalamus (HP). Six crossbred steers that grazed high-endophyte (greater than 90% infected) fescue and four steers that grazed low-endophyte (less than 1% infected) fescue from April to September were slaughtered. Brains and pituitaries were removed and dissected. Extracts from neural tissue were analyzed for DA, DOPAC and HVA using high performance liquid chromatography/electrochemical detection. Pituitary extracts and sera from blood samples taken 5 d prior to slaughter were subjected to prolactin radioimmunoassay. Consumption of high-endophyte fescue was associated with decreased concentrations of prolactin in serum (P less than .01) and in the anterior pituitary (P = .08), decreased (P less than .05) concentrations of DA in the SME and decreased (P less than .01) concentrations of HVA in the POA and HP, but it did not influence levels of DOPAC. These results suggest that endophyte toxins may reduce prolactin synthesis and release and may alter activity of dopaminergic neurons.

Evidence for endogenous opioid modulation of serum luteinizing hormone and prolactin in the steer.

Opioid modulation of LH and prolactin (PRL) concentrations in Angus steers was investigated. In Exp. 1, morphine sulfate (M) was administered at either 1, 2 or 3 mg/kg BW (n = 4) as an i.v. injection. Blood samples were obtained at 15-min intervals for 4 h pre- and post-treatment for serum hormone analyses. Mean serum LH concentration and number of LH secretory pulses decreased (P less than .1) for 2 h after M (4.1 to nadir of 2.4 ng/ml, and .33 vs. .21 pulses/h; pre- vs post-treatment). Luteinizing hormone pulse amplitude decreased (P less than .01; 7.3 vs 2.6 ng/ml; pre- vs post-treatment) during the 2 h following M. Prolactin concentrations increased 126.6%, 170.6% and 187.6% following 1, 2 and 3 mg M/kg BW, respectively (P less than .05, 1 vs 2; P less than .01, 1 vs 3). In Exp. 2, either saline solution (S, n = 6) or M (.31 mg/kg BW, i.v. injection followed by .15 mg/(kg.h) infusion; n = 6) was given for 7 h. Concentration of LH was unaffected. Response of LH to naloxone was determined in Exp. 3. Blood samples were obtained for 2 h pre- and post-administration of either naloxone (1 mg/kg BW, i.v. injection; n = 5) or S (n = 5). Response of LH at 15, 30 and 45 min posttreatment was greater (P less than .05) in naloxone- compared with S-treated steers. In summary, M had no significant effect on serum LH concentration or LH pulse frequency, but it decreased pulse amplitude and increased serum PRL concentrations. In contrast, naloxone increased LH secretion. These observations taken together indicate a physiological role for opioid modulation of LH and PRL secretion in the steer.

Influence of the ovary and suckling on luteinizing hormone response to naloxone in postpartum beef cows.

The influence of the suckling stimulus and ovarian secretions on LH response to naloxone was studied in 16 postpartum anestrous beef cows that were assigned randomly to one of four groups (n = 4/group): intact suckled (IS), intact nonsuckled (IN), ovariectomized suckled (OS) or ovariectomized nonsuckled (ON). Ovariectomy (OS + ON) and calf removal (IN + ON) were performed on d 2, 3 or 4 after parturition. Jugular venous blood was collected at 15-min intervals for 4 h before and 4 h after administration of naloxone (1 mg/kg BW, i.v.) on d 14 and d 28 after parturition. Gonadotropin-releasing hormone (5 micrograms, i.v.) was given 3 h after naloxone. Both IN and OS increased (P less than .05) mean pretreatment LH above IS values (mean +/- SE, ng/ml; IS 1.6 +/- .1 vs IN 2.5 +/- .3 and OS 2.7 +/- .4; P less than .01), whereas ON increased (P less than .01) LH (3.7 +/- .3 ng/ml) even further. Mean LH increased (P less than .05) after naloxone administration in all treatment groups. However, magnitude of this response was variable and dependent on ovarian status. Amplitude of the naloxone-induced LH response was greater (P less than .05) for ovariectomized (5.9 +/- 1.1 ng/ml) than for intact groups (2.7 +/- .5 ng/ml). Gonadotropin-releasing hormone increased mean LH concentrations in all groups. We suggest that ovarian secretions and the suckling stimulus contribute to endogenous opioid inhibition of LH during the postpartum interval.(ABSTRACT TRUNCATED AT 250 WORDS)

The bovine preoptic area and median eminence: sites of opioid inhibition of luteinizing hormone-releasing hormone secretion.

Autoradiography was used to quantify opioid receptors in the median eminence (ME) and preoptic area (POA) of the brain of eight heifers, and in vitro perifusion of ME and POA tissue from seven cows and heifers was used to examine the release of LHRH after administration of naloxone (NAL). For quantitative receptor autoradiography, [3H]NAL was used as the radioligand and NAL or morphine as competitors. Specific binding of [3H]NAL in POA and ME resulted in linear Scatchard plots with similar equilibrium dissociation constants (Kd = 4.2 +/- 1.1 nM) and mean binding site densities in the POA and ME (POA: 80.3 +/- 5.8; ME 67.5 +/- 8.0 fmol/mm2). There were no differences between mean binding site densities of zonas externa and interna of the ME; however, between various regions of the POA within individual animals, binding site densities varied threefold (47.6 to 165.1 fmol/mm2). During in vitro perifusions of isolated POA and ME, basal LHRH secretion from ME decreased (P less than .001) from 15.9 +/- 1.8 to 7.3 +/- .8 pg/10 min fraction (500 microliters) but remained constant for POA (3.1 +/- .4 pg/fraction). Injections of medium alone did not affect LHRH secretion. Although there was no significant dose (10(-9) to 10(-7) M) effect, NAL increased (P less than .05) LHRH efflux from the ME and POA when administered at 110 min from the initiation of perfusion and again at 200 min for ME but not for POA. All tissues responded to KCl (30 mM) administered at 290 min of perifusion with increased (P less than .001) LHRH efflux. Both immunoreactive-LHRH and immunoreactive-beta-endorphin were immunocytochemically localized in neurons from some of these perifused tissues. We suggest that endogenous opioids suppress LHRH secretion by actions on specific opioid receptors located within the POA and ME of the brain.

Serum prolactin and growth hormone responses to naloxone and intracerebral ventricle morphine administration in heifers.

These studies examined responses of serum prolactin (PRL) and growth hormone (GH) to opioid agonist and antagonist administration in heifers. To minimize nonspecific and behavioral effects and to facilitate future studies with specific opioid receptor agonists, a cannula was placed within the third cerebral ventricle of the brain of 4- to 10-mo-old heifers to directly access hypothalamic regions involved in the regulation of PRL and GH secretion. Increasing doses of morphine (M) from 2 to 1,500 micrograms injected into the third cerebral ventricle increased (P less than .001) serum PRL concentrations in a dose-related manner. Growth hormone responses were variable, resulting in elevated (P less than .05) serum concentrations following morphine, but no dose-related effects were apparent. Both PRL and GH responses to 700 micrograms M were absent when an intracerebral ventricle injection of an equimolar dose of naloxone, an opioid receptor antagonist, was administered prior to M. In a replicated 4 x 4 latin square, the effects of intravenous naloxone on PRL and GH responses was tested in young (86 +/- 11 d) and older (234 +/- 6 d) heifers. Naloxone at doses of 1, 2 and 4 mg/kg reduced (P less than .05) serum concentrations of PRL for 45 to 60 min. Mean concentrations of GH tended to be higher (P less than .07) in older heifers All doses of naloxone decreased (P less than .05) serum GH concentrations in older heifers but proved ineffective in younger heifers. There were no differences between doses of naloxone on either PRL or GH. These data suggest that endogenous opioids are involved in the regulation of PRL and GH secretion in heifers.

Effect of photoperiod and morphine on plasma prolactin concentrations and thyrotropin-releasing hormone secretion in the ewe.

This study investigated the effects of daily photoperiod length and morphine on thyrotropin-releasing hormone (TRH) and prolactin secretion in sheep. Two groups of adult ewes were kept under either 16 h L:8 h D or 8 h L:16 h D photoperiods for approximately 60 days. Then the photoperiods were reversed and approximately 60 days later push-pull cannulae were implanted into the hypothalamic stalk-median eminence. After a 7-day recovery period, the ewes were subjected to hypothalamic perfusion and blood was collected at 15-min intervals from the jugular vein. Perfusates also were collected into 15-min fractions. The first 3 h of perfusion served as an equilibrium period. During the next 2 h, saline was infused into one jugular vein. This was followed by a 2-hour morphine infusion (1 mg.kg-1.h-1). Data from 13 ewes were analyzed for effect of photoperiod and drug on TRH concentrations in the perfusates and prolactin in the serum. Prolactin was significantly (p less than 0.01) higher under 16 h L:8 h D than 8 h L:16 h D and was greatly increased (p less than 0.001) by morphine infusion. TRH only tended to be higher (p = 0.05) under 16 h L:8 h D than under 8 h L:16 h D, but morphine infusion induced a rapid and highly significant (p less than 0.01) increase in TRH secretion. There were no photoperiod and drug interactions for either TRH or prolactin.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteremia-induced changes in pituitary hormone release and effect of naloxone.

Acute bacteremia in sheep caused a surge of plasma beta-endorphin/beta-lipotropin (beta-EP/beta-LPH) associated with shivering behavior, tachycardia, hyperthermia, hemoconcentration, and decreased respiration rate. The surge of plasma beta-EP/beta-LPH was immediately followed by increases (P less than 0.05) in plasma prolactin and growth hormone (GH) concentrations and a depression (P less than 0.05) of plasma luteinizing hormone. These changes in pituitary hormone release were consistent with opioid-induced changes described in the literature. To examine possible opioid mediation, naloxone (2.5 mg X kg-1 X h-1) was continuously infused intravenously from 3 h before to 3 h after induction of an E. coli bacteremia. With the exception of plasma GH, naloxone failed to alter any of the hormonal or clinical parameters associated with bacteremia. For plasma GH, naloxone delayed (P less than 0.01) the increase but did not attenuate its magnitude, suggesting that an opioid mechanism may influence the timing of the pituitary GH release resulting from bacteremia. In general, opioid mechanisms sensitive to the present dosage of naloxone do not appear to mediate bacteremia-induced changes in hormonal or clinical parameters.

Continuous measurement of cerebral arteriovenous differences of beta-endorphin in sheep.

Blood was collected at 20-second intervals from the external carotid artery and from the dorsal longitudinal sagittal sinus (sagittal sinus, SS) of ovariectomized sheep. The point of SS catheterization was very near the point at which diencephalic effluent entered the SS. Concentrations of beta-endorphin (beta-EP) immunoreactivity were quantified by radioimmunoassay procedures in blood plasma and in cerebrospinal fluid (CSF) from the cisterna magna. Increases in plasma beta-EP concentration were provoked by intracarotid injection of naloxone and by experimental production of bacteremia (i.e., intravascular bacteria), but these procedures failed to increase beta-EP in CSF. Quantities of beta-EP in plasma samples from the SS were assumed to represent arterial contribution (minus tissue uptake), diencephalic secretion, and retrograde delivery of pituitary beta-EP to the diencephalic effluent. The arterial contribution was removed mathematically by subtracting the arterial plasma beta-EP concentration from the concurrent SS plasma concentration of beta-EP to yield a paired arteriovenous (AV) difference. When this AV difference was consistently positive and satisfied our statistical criterion for being greater than zero, we concluded that either pituitary beta-EP was delivered in a retrograde manner to diencephalon or the diencephalon secreted beta-EP. However, this situation occurred in only 5 of 31 periods examined. Furthermore, only 2 of these 5 periods occurred during times of increasing arterial concentrations of beta-EP. Such concurrence would be expected if both changes were caused by a major discharge of beta-EP from the pituitary gland. Therefore, the present results provide little evidence for retrograde delivery of pituitary beta-EP to the brain without systemic dilution.

Effect of frontal hypothalamic deafferentation on duration of breeding season and melatonin secretion in the ewe.

The objectives of this study were to determine if ewes subjected to frontal hypothalamic deafferentation (FHD) during anestrus remained anestrus or began to have estrous cycles, and if melatonin secretion was disrupted by FHD. Ovary-intact ewes in Group 1 were subjected to either FHD (n = 10) or sham FHD (n = 5) in early July 1983. Estrous cycles were monitored by measuring circulating progesterone concentrations from before FHD until September 1985. Group 2 ewes (n = 4) were subjected to FHD in October 1984. In late April 1985, blood samples were taken from all ewes at 1- to 4-h intervals from 1100 h to 0700 h of the following day to monitor diurnal changes of melatonin. Hypothalami were collected for histological evaluation of lesions. All Group 1 ewes (sham FHD and FHD) initiated normal estrous cycles in August and September 1983, and all ceased cycles by mid-February 1984. All sham FHD and 4 FHD ewes remained anestrus until August or September of 1984 and then resumed normal cycles. In contrast, 5 FHD ewes resumed cycles as early as April 1984 and then cycled intermittently or almost continuously. Two Group 2 ewes cycled continuously after FHD and 2 cycled infrequently. FHD ewes that showed prolonged breeding seasons had cuts that damaged the suprachiasmatic nucleus (SCN) and adjacent structures. Mean nocturnal (2000 h-0500 h) melatonin concentrations did not differ (p greater than 0.05) between sham FHD, FHD "normal season," and FHD "continuous cycle" ewes. In summary, damage to the SCN region by FHD during anestrus had no detectable effect on either onset or cessation of the next breeding season but greatly prolonged subsequent breeding seasons. Thus, the environmental signals that both initiated and terminated the 1983 breeding season apparently had been given before FHD was performed in midsummer. Damage to the SCN region during the breeding season caused some ewes to cycle continuously. The effects of FHD apparently were not due to disruption of melatonin secretion. FHD ewes that showed prolonged breeding seasons had normal seasonal changes of plasma prolactin concentrations. This suggests that different neural structures control seasonal patterns of gonadotropin and prolactin secretion.

Radioimmunological measurement of beta-endorphin in equine plasma.

Radioimmunoassay procedures were developed and validated for the quantification of beta-endorphin (beta-EP)-like immunoreactivity in equine plasma. beta-EP could be quantitatively extracted from plasma with silicic acid powder and subsequently assayed, however, valid estimates of this hormone could also be obtained on unextracted plasma. Although beta-lipotropin (beta-LPH) cross-reacted in the assay, it was not necessary to correct for beta-LPH activity when assaying unextracted plasma because chromatographic analyses showed that 92% of the immunoreactivity in plasma extracts was similar in molecular weight to authentic beta-EP (1-31). In addition, electroacupuncture treatment did not alter the relative proportion of immunoreactivity among different molecular weight fractions.