[Effect of electroacupuncture on pain threshold and expression of pain-related factors cyclooxygenase-2, prostaglandin E2 and ß-endorphin in rats with chronic pelvic pain syndrome].

OBJECTIVE: To explore the possible mechanism of electroacupuncture (EA) underlying improvement of chronic pelvic pain syndrome (CPPS). METHODS: Fifty SD rats were randomly divided into control, model, sham operation, EA and sham EA groups (n=10 rats in each group). The CPPS model was established by injecting complete Freund's adjuvant (CFA, 50 µL) into the ventral lobes of the prostate. EA (2 Hz/100 Hz) was applied to "Guanyuan"(CV4), "Zhongji"(CV3), "Sanyinjiao" (SP6) and "Huiyang"(BL35) once daily for 40 min, 5 days a week for 4 weeks, while rats in the sham EA group were treated with the same acupoints but without electrical stimulation. Mechanical pain threshold (MPT) and heat pain threshold (HPT) were measured before and after intervention. The body weight and prostate weight were measured and prostate index was calculated. Histopathological changes of prostate tissue were observed by HE staining. The levels of cycloxygenase-2 (COX-2), prostaglandin E2 (PGE2) and ß-endorphin (ß-EP) in prostate tissue were detected by ELISA. RESULTS: Compared with the control and sham operation groups, the MPT and HPT were significantly lower (P<0.01), and the prostate weight, prostate index, the contents of PGE2 and COX-2 were significantly increased (P<0.01), while the content of ß-EP was significantly decreased (P<0.01) in the model group. Compared with the model group, the MPT and HPT were significantly increased (P<0.01) after 3 and 4 courses of treatment, and the prostate weight, prostate index, the contents of PGE2 and COX-2 were significantly decreased (P<0.01), while the content of ß-EP was significantly increased (P<0.01) in the EA group, rather than in the sham EA group (P>0.05). CONCLUSION: EA can effectively relieve pain in CPPS rats, which may be related to its functions in down-regulating COX-2 and PGE2, and up-regulating ß-EP.

[Effect of Electroacupuncture Intervention on ß-endorphin Levels of Hypothalamus and Spinal Cord Tissues in Rats with Pelmatic Incisional Pain].

OBJECTIVE: To observe the effect of electroacupuncture (EA) intervention on pain thresholds (PT) and contents of ß-endorphin (EP) in the hypothalamus and spinal cord, and the expression of 5-HT in the dorsal raphe nucleus(DRN)in rats with pelmatic incisional pain, so as to investigate the underlying mechanisms of acupuncture in reducing post-operative pain. METHODS: Wistar rats were randomized into normal control, model, EA and non-acupoint groups (n=8/group). The pelmatic pain model was induced by making an incision (about 1 cm in length, to the fascia and muscle layers) from the heel towards the toes. EA (2 Hz, 1.5-2 V) was applied to "Zusanli" (ST 36) and "Kunlun" (BL 60) or non-acupoint (about 3 mm beside the ST 36 and BL 60) on the affected side for 20 min, once daily for three days. The thermal PT and mechanical PT were measured before and after operation and after EA. The contents of ß-EP in hypothalamus and L3-S4 spinal cord were detected using enzyme linked immunosorbent assay (ELISA) and the expressions of ß-EP in hypothalamus and 5-HT in DRN were measured with immunohistochemistry. RESULTS: After EA intervention, the markedly decreased mechanical and thermal pain thresholds on day 1 and 3 after paw incision were significantly increased in the EA group (P<0.05), but not in the non-acupoint group (P>0.05). The hypothalamic ß-EP content was significantly higher in the model group than in the normal group (P<0.05), and further up-regulated in the EA group (not the non-acupoint group) than in the model group (P<0.05). In addition, the hypothalamic ß-EP immunoreactive (IR)-positive cell number and 5-HT immunoactivity level in DRN were also considerably up-regulated in the EA group (P<0.05) but not in the non-acupoint group (P>0.05). No significant changes were found in the lumbar spinal ß-EP contents in the model, EA and non-acupoint groups (P>0.05). CONCLUSIONS: EA stimulation of "Zusanli"(ST 36) and "Kunlun" (BL 60) has an analgesic effect in pelmatic incision pain rats, which may be related to its effects in raising the level of hypothalamic ß-EP and the expression of 5-HT in DRN.

Neuronal responses in the brainstem and hypothalamic nuclei following insulin treatment during the late follicular phase in the ewe.

The aim of this study was to determine the neuronal responses following insulin administration during the late follicular phase. Intact ewes were given either saline or insulin (5 IU/kg, i.v.) at 35 h after progesterone withdrawal and killed 3 h later. There was a marked increase in the number of Fos-positive noradrenergic neurones in the caudal brainstem of insulin-treated ewes. In the hypothalamic paraventricular nucleus, insulin treatment increased the presence of Fos-positive corticotrophin-releasing hormone neurones (from 2% to 98%) and Fos-positive arginine vasopressin parvocellular neurones (from 2% to 46%). Interestingly, after insulin treatment, despite a general increase in Fos-positive neurones in the arcuate nucleus (ARC), there was a marked reduction (from 47% to 1%) in Fos-positive ß-endorphin neurones. Similarly, colocalized Fos and oestradiol receptor (ER) α-positive neurones decreased in the ARC after insulin (from 7% to 3%). Conversely, in the ventromedial nucleus, ERα-positive neurones with Fos increased (from 7% to 22%) alongside a general increase in Fos-positive neurones. Overall, a complex system of neurones in brainstem and hypothalamus is activated following insulin administration during the late follicular phase.

Protective effect of electroacupuncture at the Neiguan point in a rabbit model of myocardial ischemia-reperfusion injury.

OBJECTIVES: To investigate possible cardioprotective mechanisms of electroacupuncture (EA) at the Neiguan point and at the Lieque point in the presence of myocardial ischemia-reperfusion injury (MIRI). METHODS: The changes in ventricular tissue Bax and Bcl-2 protein expression, malondialdehyde (MDA) content and glutathione peroxidase (GSH-PX) activity were examined, as well as beta-endorphin (beta-EP) con-tent in a rabbit model of MIRI. Four randomized groups were studied: sham, untreated MIRI, MIRI followed by EA at the Neiguan point, and MIRI followed by EA at the Lieque point. The MIRI model involved ligating the left anterior descending coronary artery for 30 min followed by a 60 min postischemia reperfusion period. RESULTS: EA at the Neiguan point dramatically decreased the number of apoptotic cells and the content of beta-EP and MDA, and inhibited Bax protein expression while enhancing Bcl-2 expression and GSH-PX activity. Furthermore, EA enhanced Bcl-2 expression and GSH-PX activity. Lesser effects were elicited by EA at the Lieque point. CONCLUSIONS: The cardioprotective effects of applying EA at the Neiguan point on MIRI include reducing apoptosis, regulating apoptosis- controlling genes, and decreasing myocardial MDA and beta-EP while enhancing GSH-PX activity.

Brain-region responsiveness to DT56a (Femarelle) administration on allopregnanolone and opioid content in ovariectomized rats.

OBJECTIVE: The natural selective estrogen receptor modulator DT56a (Femarelle), derived from soybean, has been shown to relieve menopausal vasomotor symptoms with no effect on sex steroid hormone levels or endometrial thickness.The purpose of the present study was to evaluate the neuroendocrine effect of DT56a administration through the evaluation of brain content of allopregnanolone (AP), an endogenous neurosteroid gamma-aminobutyric acid agonist with anxiolytic properties, and through the assessment of beta-endorphin (beta-END), the endogenous opioid implicated in pain mechanism, emotional state, and autonomic control. METHODS: Five groups of Wistar ovariectomized (OVX) rats received one of the following treatments: oral DT56a administration at doses of 6, 12, 60, and 120 mg kg(-1) day(-1) or estradiol valerate (E2V) at a dose of 0.05 mg kg(-1) day(-1) for 14 days. One group of fertile and one group of OVX rats receiving placebo were used as controls. The concentration of AP was assessed in the frontal and parietal cortex, hippocampus, hypothalamus, anterior pituitary, and serum, whereas the content of beta-END was evaluated in the frontal and parietal cortex, hippocampus, hypothalamus, neurointermediate lobe, anterior pituitary, and plasma. RESULTS: DT56a increased AP levels in all brain areas analyzed and in serum, with a classical dose-related curve in comparison with OVX rats. In some brain areas, such as the frontal cortex, the parietal cortex, and the anterior pituitary, positive results were found even with the administration of a lower DT56a dose of 60 mg kg(-1) day(-1), attaining AP levels in the range of those in animals treated with E2V. Similarly, beta-END levels were enhanced in selected brain areas such as the hippocampus, the hypothalamus, the neurointermediate lobe, and the anterior pituitary in comparison with those in OVX rats, in which the increase of the opioid was dose related and in the range of those in rats treated with E2V. CONCLUSIONS: This study demonstrated that DT56a positively affects brain neurosteroidogenesis and the opiatergic system: DT56a exerts an estrogen-like effect on selective areas related to mood, cognition, and homeostasis control, presenting a specific pattern of interaction with the brain function. These findings may, in part, explain the clinical effect of DT56a on menopausal symptoms.

[Effects of electroacupuncture on plasma and cerebral somatostatin and beta-EP contents and learning-memory ability in vascular dementia rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on learning-memory ability of vascular dementia (VD) rats, and the simultaneous changes of plasma and cerebral somatostatin (SS) and beta-endorphin (EP) contents. METHODS: Forty-one SD rats were randomly divided into control (n=8), model A (n=8, no treatment) and B (n=8, intragastric perfusion of 15% saline), EA (n=9) and medication (n=8, intragastric perfusion of Nimoldipine, 12 mg/kg) groups. VD model was established by using modified 4-vessels occlusion method. EA (150 Hz, 1-2 mA) was applied to "Baihui" (GV 20), "Dazhui" (DU 14), "Pishu" (BL 20) and "Shenshu" (BL 23) for 20 min, once daily for 15 days. Morris water maze tests were conducted for evaluating the rats' learning-memory ability. The contents of SS and beta-EP in plasma and brain tissue were measured by radioimmunoassay (RIA). RESULTS: In comparison with sham-operation group, the escape latency (EL) prolonged significantly and the target-platform crossing times decreased remarkably (P<0.01) in model group B. In comparison with model group B, EL shortened and target-platform crossing times increased both significantly in EA and medication groups (P<0.01). Plasma and cerebral SS, and cerebral beta-EP contents of model groups A and B were significantly lower than those of sham-operation group(P<0.01), while plasma beta-EP level had no obvious change (P>0.05). Plasma and cerebral SS, and cerebral beta-EP contents in both EA and medication groups were considerably higher than those in model groups A and B (P<0.01). No significant differences were found between EA and medication groups in EL, target-platform crossing times, plasma and cerebral SS and beta-EP levels, and between model group A and model group B in plasma and cerebral SS and beta-EP levels (P>0.05). CONCLUSION: EA can raise plasma and cerebral SS and cerebral beta-EP levels, and improve the learning-memory ability in VD rats.

[Influence of connexin 43 gene knockout on the analgesic effect of acupuncture in visceral pain mice].

OBJECTIVE: To explore the possible central and peripheral mechanisms of connexin 43 gene knockout in acupuncture analgesia. METHODS: Fifty-four wide type (WT) mice and 54 connexin 43 gene knockout (heterozygote, HT) mice were randomly divided into WT control group, WT model group, WT acupuncture group, HT control group, HT model group and HT acupuncture group (n = 18/group). Visceral pain model was established by intraperitoneal injection of 0.6% acetic acid (0. 1 mL/10 g). "Zhongwan" (CV 12) and bilateral "Zusanli" (ST 36) were punctured with filiform needles and stimulated for 30 min by manipulating the needle for 30 s every 5 min. The latency and the number of body-writhing response were observed and the contents of beta-endorphin (beta-EP) in hypothalamus and serum prostaglandin E2 (PGE2) were detected with radioimmunoassay (RIA). RESULTS: There was no significant difference in the latency and the number of body-writhing response, and contents of hypothalamic beta-EP and serum PGE2 between HT and WT control groups (P > 0.05). Compared with the corresponding control groups, the latency of body-writhing in WT model and HT model groups shortened significantly and the number of body- writhing increased considerably in two model groups (P < 0.01). While in comparison with WT model group, the latency of body-writhing prolonged significantly and the number of body-writhing decreased apparently in WT acupuncture group (P < 0.01); but no marked differences were found between HT model and HT acupuncture groups in these two indexes (P > 0.05). Compared with the corresponding control groups, the contents of both beta-EP and PGE2 increased obviously in WT model and HT model groups (P < 0.05). In comparison with WT model group, beta-EP levels in WT acupuncture group increased further significantly (P < 0.05), and serum PGE2 in WT acupuncture group decreased obviously (P < 0.05); but no significant changes were found between HT model and HT acupuncture groups in beta-EP and PGE2 levels (P > 0.05). CONCLUSION: Acupuncture has a marked antinociceptive effect in WT mice with visceral pain, which may be related to its effects in increasing hypothalamic beta-EP and decreasing serum PGE2; while in connexin 43 gene knockout mice, all the above-mentioned effects of acupuncture are eliminated, indicating an important role of connexin 43 in the analgesic effect of acupuncture.

beta-Endorphin concentration in colostrums of Burkinabe and Sicilian women.

OBJECTIVE: Endorphins (EPs) present in human colostrum may be relevant for immediate postnatal fetal adaptation because this compound is involved in stress response and adaptation mechanisms. Endorphin levels in human colostrum are two-fold greater than corresponding maternal plasma levels; however, the high endorphin levels in human milk decrease as lactation continues. The aim of this study was to determine the beta-EP concentration in colostrums of women residing in Burkina Faso and Sicily. In addition, we investigated the source of potential differences in beta-EP levels between these populations, especially ethnic sources of these deviations. METHODS: The concentration of beta-EP was determined in the colostrum from the first 3 d subsequent to delivery by an enzyme immunoassay as immunoreactive material (IRM). RESULTS: The production of beta-EP in the colostrum was significantly higher in Burkinabe mothers (0.83 +/- 0.04 ng/mL) than in Sicilian mothers (0.31 +/- 0.02 ng/mL) at 24 h after delivery. Colostrum levels of beta-EP declined progressively during the first 3 d after delivery in both populations (0.64 +/- 0.1 and 0.28 +/- 0.015 ng/mL, respectively, at 72 h). The level of beta-EP-IRM correlated significantly with pain and psychological involvement during and after delivery. In addition, the correlation between beta-EP-IRM and length of stage II of labor was significant (P < 0.0001) in the colostrums of Sicilian mothers who received ergot derivatives, episiorrhaphy, and child birth preparation. The correlation between beta-EP-IRM and length of stage II was less significant (P < 0.001) in the colostrums of Burkinabe mothers who received neither ergot derivatives nor child birth preparation. CONCLUSION: During the first 3 d after labor the beta-EP-IRM concentration in the colostrums of Burkinabe mothers differs from that of Sicilians. In addition, because Burkinabe women produce a larger volume of colostrum, their newborns receive, during the first days of life, a larger absolute amount of beta-EP-IRM, likely resulting in better postnatal fetal adaptation.

Beta-endorphin, Met-enkephalin and corresponding opioid receptors within synovium of patients with joint trauma, osteoarthritis and rheumatoid arthritis.

OBJECTIVE: Intra-articularly applied opioid agonists or antagonists modulate pain after knee surgery and in chronic arthritis. Therefore, the expression of beta-endorphin (END), Met-enkephalin (ENK), and mu and delta opioid receptors (ORs) within synovium of patients with joint trauma (JT), osteoarthritis (OA) and rheumatoid arthritis (RA) were examined. METHODS: Synovial samples were subjected to double immunohistochemical analysis of opioid peptides with immune cell markers, and of ORs with the neuronal markers calcitonin gene-related peptide (CGRP) and tyrosine hydroxylase (TH). RESULTS: END and ENK were expressed by macrophage-like (CD68(+)) and fibroblast-like (CD68(-)) cells within synovial lining layers of all disorders. In the sublining layers, END and ENK were mostly expressed by granulocytes in patients with JT, and by macrophages/monocytes, lymphocytes and plasma cells in those with OA and RA. Overall, END- and ENK-immunoreactive (IR) cells were more abundant in patients with RA than in those with OA and JT. ORs were found on nerve fibres and immune cells in all patients. OR-IR nerve fibres were significantly more abundant in patients with RA than in those with OA and JT. muORs and deltaORs were coexpressed with CGRP but not with TH. CONCLUSIONS: Parallel to the severity of inflammation, END and ENK in immune cells and their receptors on sensory nerve terminals are more abundant in patients with RA than in those with JT and OA. These findings are consistent with the notion that, with prolonged and enhanced inflammation, the immune and peripheral nervous systems upregulate sensory nerves expressing ORs and their ligands to counterbalance pain and inflammation.

Methodological aspects of rat beta-endorphin analysis-influence of diurnal variation.

Beta-endorphin radioimmunoassays (RIAs) are widely performed following physical, emotional and environmental challenges in the rat. In the literature, a wide range of techniques have been described, but in the present study, we have focused on methodological aspects of beta-endorphin RIAs, investigating various characteristics of human and rat specific antibodies. Initial studies verified that the RIA outcome was not appropriate when using non-species compatible components. Novel rat beta-endorphin antibodies, r 4114 and r 4268, were raised in rabbits and characterised in terms of specificity, avidity and titer. Both of the new antisera showed 68.1% cross-reactivity with human beta-endorphin. The ED50 was 50+/-8 pmol/l, and the mean ED80 was 17 pmol/l for r 4268 but three-fold higher for r 4114. The intra-assay coefficient of variation (CV) was 7% at 100 pmol/l and the inter-assay CV was 10% at the same level for r 4268 and similar for r 4114. Using this novel rat beta-endorphin RIA for analyses of diurnal influence and removal from the Animal House cage, no significant changes were observed in either the hypothalamus or peri-aqueductal grey regions. These results suggest that rat beta-endorphin concentrations in these brain areas are not affected by order of removal or diurnal variation.

Effect of tibolone administration on central and peripheral levels of allopregnanolone and beta-endorphin in female rats.

OBJECTIVE: We evaluated the effects of tibolone oral administration on neuroendocrine function by investigating the modulation exerted by tibolone administration on allopregnanolone and central and peripheral beta-endorphin (beta-EP) levels in ovariectomized rats. DESIGN: Female Wistar rats (N = 64) were included: 48 rats were ovariectomized, 8 cycling rats were included as controls, and 8 cycling rats were treated with placebo. The ovariectomized animals were divided into six groups: untreated rats and those that received 14-day oral treatment with either placebo, estradiol valerate (E2V) 0.05 mg/kg/d, or tibolone (0.1, 0.5, or 2 mg/kg/d. beta-EP levels were assessed in the frontal lobe, parietal lobe, hippocampus, hypothalamus, anterior pituitary, neurointermediate pituitary, and plasma, whereas allopregnanolone levels were measured in the frontal lobe, parietal lobe, hippocampus, hypothalamus, anterior pituitary, adrenal glands, and serum. RESULTS: The administration of tibolone (0.5 and 2 mg/kg/d) in ovariectomized rats induces a significant increase of allopregnanolone in the frontal lobe, parietal lobe, hippocampus, hypothalamus, whereas in serum a significant increase of allopregnanolone occurs only with the dose of 2 mg/kg/d, a significant decrease in allopregnanolone levels occurs in the adrenal glands. No changes occurred in the anterior pituitary. Tibolone doses of 0.5 and 2 mg/kg/d induced a significant increase in beta-EP content in the frontal lobe, hypothalamus, and neurointermediate lobe; and, at doses of 2 mg/kg/d, in the parietal lobe, anterior pituitary, and plasma, without changes in the hippocampus. Compared with E2V, 0.5 mg/kg/d tibolone showed a similar effect on allopregnanolone and beta-EP in most brain regions. CONCLUSIONS: Tibolone administration affects beta-EP and allopregnanolone levels, playing a role as a neuroendocrine modulator.

Only arginine vasopressin, not oxytocin and endogenous opiate peptides, in hypothalamic paraventricular nucleus play a role in acupuncture analgesia in the rat.

Our previous study proved that hypothalamic paraventricular nucleus (PVH) plays an important role in acupuncture analgesia. The effect of acupuncture on the concentrations of arginine vasopressin (AVP), oxytocin (OXT), leucine-enkephaline (L-Ek), beta-endorphin (beta-Ep) and dynorphinA(1-13) (DynA(1-13)) was investigated in rat PVH. Electrical acupuncture of "Zusanli" points (St. 36) 30 min increased the AVP, not OXT, L-Ek, beta-Ep and DynA(1-13) concentrations in PVH tissue using micropunch and radioimmunoassay, which showed a negative relationship between the pain threshold and AVP concentrations in PVH tissue. Electrical acupuncture could elevate the AVP concentrations in PVH perfuse liquid during acupuncture, and then reduce the AVP concentrations in PVH perfuse liquid after acupuncture. But no change in OXT, L-Ek, beta-Ep and DynA(1-13) concentrations was detected in PVH perfuse liquid. Electrical acupuncture decreased the number of AVP, not OXT, L-Ek, beta-Ep and DynA(1-13) immunoreactive cells in PVH using immunocytochemistry. The results suggested that only AVP, not OXT and endogenous opiate peptides in PVH involved acupuncture analgesia in the rat.

Age-related changes in the oestrous cycle and reproductive hormones in senescence-accelerated mouse.

To investigate age-related changes in the oestrous cycle and reproductive hormone levels in senescence-accelerated mouse (SAM), we examined these parameters in 3-, 5-, 7-, 9- and 11-month-old female SAM-prone/8 (SAMP8) and SAM-resistant/1 (SAMR1) strains. Levels of beta-endorphin (beta-EP) and substance P (SP) in the hypothalamus were also measured. The oestrous cycle and dioestrus of 9-month-old SAMP8 mice were significantly prolonged compared with age-matched SAMR1 mice. Furthermore, the concentration of serum oestradiol was lower and the level of pituitary luteinising hormone was higher in SAMP8 mice compared with SAMR1 mice. This characterises the hypothalamus-pituitary-ovary (HPO) axis of the SAMP8 strain as hypergonadotropic-hypogonad. The levels of beta-EP and SP in the SAMP8 hypothalamus were lower than in the SAMR1 hypothalamus. These results indicate that the function of the HPO axis in SAMP8 mice declines early and this may be attributed, in part, to the decline in beta-EP and SP concentrations in the hypothalamus.

Obliteration of alpha-melanocyte-stimulating hormone derived from POMC in pituitary and brains of PC2-deficient mice.

Alpha-melanocyte-stimulating hormone (alpha-MSH) is a neuropeptide expressed in pituitary and brain that is known to regulate energy balance, appetite control, and neuroimmune functions. The biosynthesis of alpha-MSH requires proteolytic processing of the proopiomelanocortin (POMC) precursor. Therefore, this study investigated the in vivo role of the prohormone convertase 2 (PC2) processing enzyme for production of alpha-MSH in PC2-deficient mice. Specific detection of alpha-MSH utilized radioimmunoassay (RIA) that does not crossreact with the POMC precursor, and which does not crossreact with other adrenocorticotropin hormone (ACTH) and beta-endorphin peptide products derived from POMC. alpha-MSH in PC2-deficient mice was essentially obliterated in pituitary, hypothalamus, cortex, and other brain regions (collectively), compared to wild-type controls. These results demonstrate the critical requirement of PC2 for the production of alpha-MSH. The absence of alpha-MSH was accompanied by accumulation of ACTH, ACTH-containing imtermediates, and POMC precursor. ACTH was increased in pituitary and hypothalamus of PC2-deficient mice, evaluated by RIA and reversed-phase high pressure liquid chromatography (RP-HPLC). Accumulation of ACTH demonstrates its role as a PC2 substrate that can be converted for alpha-MSH production. Further analyses of POMC-derived intermediates in pituitary, conducted by denaturing western blot conditions, showed accumulation of ACTH-containing intermediates in pituitaries of PC2-deficient mice, which implicate participation of such intermediates as PC2 substrates. Moreover, accumulation of POMC was observed in PC2-deficient mice by western blots with anti-ACTH and anti-beta-endorphin. In addition, increased beta-endorphin1-31 was observed in pituitary and hypothalamus of PC2-deficient mice, suggesting beta-endorphin1-31 as a substrate for PC2 in these tissues. Overall, these studies demonstrated that the PC2 processing enzyme is critical for the in vivo production of alpha-MSH in pituitary and brain.

Estradiol valerate and alcohol intake: a comparison between Wistar and Lewis rats and the putative role of endorphins.

Studies show that estrogens can influence alcohol consumption; however, findings are variable and an etiology remains unknown. Furthermore, estrogen administration can alter several neurotransmitter systems implicated in alcohol consumption, including the beta-endorphin (beta-EP) system. The present studies investigate (a) whether estradiol valerate (EV) alters voluntary alcohol consumption in Wistar and Lewis rats, (b) if an effect of EV on drinking is associated with changes in hypothalamic or pituitary beta-EP content, and (c) whether differences in alcohol drinking between treatment and rat groups are related to locomotor or defensive behavior/anxiety scores. Of 30 Wistar and 30 Lewis rats used in this study, half were injected with 2 mg EV in 0.2 ml sesame oil, while the remainder were injected with the vehicle only. After 8 weeks, all animals were tested in the open field and elevated plus maze. A week later, 4-6 animals in each group were sacrificed. The remaining animals were tested for voluntary alcohol drinking for 24 days prior to being sacrificed on the last day. Radioimmunoassay was used to estimate hypothalamic and pituitary beta-EP content. Wistar and Lewis rats injected with EV showed an increase in alcohol drinking, but their behavior scores and beta-EP levels remained unaltered. This result suggests that any EV effect on drinking is unrelated to changes in beta-EP or behavioral performance. Furthermore, Wistar rats show higher alcohol drinking, locomotor and defensive behavior scores, and hypothalamic beta-EP than Lewis rats. Higher alcohol drinking by Wistar rats might be due to higher behavioral scores or endogenous opioid activity/sensitivity.

Nucleus accumbens beta-endorphin levels are not elevated by brain stimulation reward but do increase with extinction.

Beta-endorphin is an endogenous opioid peptide implicated in reward processes, but the brain sites directly involved in its putative role in reward have not been identified. Here we used in vivo microdialysis in rats to study the effect of a potent reinforcer, lateral hypothalamus self-stimulation (LHSS), on the extracellular levels of beta-endorphin in the nucleus accumbens (NAS). The NAS is involved in the reinforcing effects of natural and artificial rewards, has high density of opioid receptors and is innervated by arcuate nucleus beta-endorphin neurons. LHSS had no effect on extracellular levels of beta-endorphin in the NAS. Surprisingly, extinction of the self-stimulation behaviour induced a rapid increase in NAS beta-endorphin levels. In a subsequent experiment in rats previously trained to self-administer heroin for 10 days, beta-endorphin levels also were increased during a test for extinction of the heroin-reinforced behaviour. Finally, the increase in extracellular beta-endorphin levels in the NAS was also observed during exposure to an aversive stimulus, intermittent footshock (20 min). These results indicate a possible role for increased levels of NAS beta-endorphin in the organism's adaptive response to stress and frustration.

Galanin modulates the activity of proopiomelanocortin neurons in the isolated mediobasal hypothalamus of the male rat.

It has become apparent that galanin as well as proopiomelanocortin-derived peptides, such as beta-endorphin, play an important role in the hypothalamic circuitry that regulates neuroendocrine functions and appetite behavior. We have recently shown that GalR1 and GalR2 galanin receptor mRNAs are expressed in proopiomelanocortin neurons of the arcuate nucleus, suggesting a direct modulatory action of galanin on the proopiomelanocortin neuronal system. In the present study, we investigated the effect of galanin on beta-endorphin release and proopiomelanocortin mRNA expression from male rat mediobasal hypothalamic fragments incubated ex vivo. Galanin induced a decrease of spontaneous beta-endorphin release within the first 30-60 min of incubation and this effect was blocked by the galanin receptor antagonist galantide. Co-incubation of galanin with FK-506 (tacrolimus), a calcineurin inhibitor, suppressed the inhibitory effect of galanin on beta-endorphin release, suggesting that calcineurin is involved in the galanin-evoked decrease in beta-endorphin release. Measurement of beta-endorphin levels in the tissues at the end of the incubation period (120 min) revealed that galanin caused a two-fold increase of beta-endorphin peptide concentration in the mediobasal hypothalamic tissues. Concurrently, galanin induced an increase in the mean density of silver grains overlying proopiomelanocortin neurons after 60 min of incubation, an effect antagonized by galantide. Finally, reverse transcription-polymerase chain reaction analysis revealed that the mRNAs for the three galanin receptor subtypes (i.e. GalR1, GalR2, and GalR3) were expressed in the incubated mediobasal hypothalamic fragments. Taken as a whole, our results indicate that galanin plays a modulatory role on proopiomelanocortin neurons and this interrelation contributes to the elucidation of the neural circuitry that controls, among others, gonadotropin-releasing hormone function.

Beta endorphin concentrations in human milk.

BACKGROUND: The source and regulatory mechanisms that elevate beta-endorphin (beta-EP) approximately twofold higher than circulating plasma levels in the colostrum of lactating mothers are still unknown, and no studies have examined beta-EP availability previously during maturation phases of human milk. Therefore, the aim of this study was to determine whether concentrations of beta-EP vary over time between colostrum, transitional, and mature breast-milk and to evaluate whether this depends on the method of delivery. METHODS: Mothers of healthy full-term and pre-term newborn infants who planned to breast-feed their newborn infants were considered for this study. They were consecutively recruited in one of 3 groups of 14, according to delivery method: group 1, vaginal delivery at term (gestational age 40.2 +/- 0.3 weeks; birth weight, 3.48 +/- 0.09 kg); group 2, preterm vaginal delivery (gestational age, 35.6 +/- 0.3 weeks; birth weight, 2.49 +/- 0.08 kg); and group 3, at-term elective cesarean section (gestational age, 39.0 +/- 0.3 weeks; birth weight, 3.32 +/- 0.14 kg). Three consecutive breast milk samples were obtained on the fourth day after birth, before each mother's discharge, and thereafter on the 10th and 30th postpartum days, close to expression of the colostrum, transitional, and mature milk production phases, respectively, to test beta-EP concentrations (beta-Endorphin 125I RIA; INCSTAR Corporation, Stillwater, MN). Data are presented as mean +/- standard deviation. Statistical comparison of beta-EP concentration among the three lactating mother groups was performed using the Kruskal-Wallis nonparametric test. In addition, to test the hypothesis of a trend toward smaller values with time of beta-EP, the authors computed within each mother group a P value per trend (Kruskal-Wallis test) of beta-EP concentration averages on the 4th, 10th, and 30th days, respectively. Student's t test for independent samples was used for the analysis of the other data. The 0.05 significance level was used in the statistical analysis. All computations were made by computer. RESULTS: Colostrum beta-EP concentrations on the fourth postpartum day of group 1 and group 2 mothers who were delivered of a neonate vaginally, at term, or prematurely were significantly higher (P < 0.01) than colostrum levels of group 3 mothers who underwent cesarean section. Group 2 mothers who were delivered of a neonate vaginally and prematurely presented the highest beta-EP concentrations (P < 0.05), lasting until the transitional milk phase (10th day). No significant differences were found across all 3 groups of lactating mothers in mature milk (30th day) beta-EP concentrations. In addition, the beta-EP trend toward smaller values with time within each of the three groups on days 4, 10, and 30 was statistically significant (P < 0.01 per trend). CONCLUSIONS: It is hypothesized that elevated beta-EP concentrations in colostrum and transitional milk of mothers who were vaginally delivered of infants may contribute to postnatal fetal adaptation, to overcoming birth stress of natural labor and delivery, and at the same time to the postnatal development of several related biologic functions of breast-fed infants.

Labor pain effects on colostral milk beta-endorphin concentrations of lactating mothers.

beta-Endorphin (beta-EP) levels of colostral milk are approximately two-fold higher than in plasma of lactating women, in who concentrations peak at term, after the first and second stages of labor. We investigated the effect of labor pain and vaginal parturition on colostral beta-EP concentrations (beta-endorphin (125)I RIA, Incstar Corp., Stillwater, Minn., USA) of at-term nursing mother, in comparison to those having undergone elective cesarean section. Our results show that colostral milk beta-EP concentrations of mothers who delivered vaginally are significantly higher on the 4th postpartum day (6.0 +/- 0.5 vs. 4.3 +/- 0.4 pmol/l, respectively; p < 0.01) than colostral levels of mothers who underwent cesarean section. These data indicate the important influence of the labor process on the colostral opioid galactopoiesis. It is suggested that labor and parturition pain may increase colostral milk beta-EP concentrations of lactating mothers in order to help the newborn overcome the stressful perinatal events of natural labor and delivery.