Proopiomelanocortin-producing neurons in the arcuate nucleus of the hypothalamus secrete beta-endorphin (beta-EP), which controls varieties of body functions including the feedback regulation of the CRH neuronal activity in the paraventricular nucleus of the hypothalamus. Whether ethanol exposure in developing rats induces beta-EP neuronal death and alters their influence on CRH neurons in vivo has not been determined. We report here that binge-like ethanol exposures in newborn rats increased the number of apoptotic beta-EP neurons in the arcuate nucleus of the hypothalamus. We also found that immediately after ethanol treatments there was a significant reduction in the expression of proopiomelanocortin and adenylyl cyclases mRNA and an increased expression of several TGF-beta1-linked apoptotic genes in beta-EP neurons isolated by laser-captured microdissection from arcuate nuclei of young rats. Several weeks after the ethanol treatment, we detected a reduction in the number of beta-EP neuronal perikarya in arcuate nuclei and in the number of beta-EP neuronal terminals in paraventricular nuclei of the hypothalamus in the treated rats. Additionally, these rats showed increased response of the hypothalamic CRH mRNA to the lipopolysaccharide challenge. The ethanol-treated animals also showed incompetent ability to respond to exogenous beta-EP to alter the lipopolysaccharide-induced CRH mRNA levels. These data suggest that ethanol exposure during the developmental period causes beta-EP neuronal death by cellular mechanisms involving the suppression of cyclic AMP production and activation of TGF-beta1-linked apoptotic signaling and produces long-term structural and functional deficiency of beta-EP neurons in the hypothalamus.

The role of β-endorphin (β-EP) in ethanol-altered NK cell cytolytic activity is studied using male Fischer-344 rats as an animal model. Ethanol was administered for 1, 2, 3, or 4 wk in a liquid diet containing 8.7% ethanol (v/v), which means that 37% of the total calories were derived from ethanol. Rats treated with ethanol for 1 wk showed an increase in hypothalamic and plasma levels of immunoreactive (IR)-β-EP, but displayed no significant effect on NK cell activity determined by 51Cr release assay, as compared with those in pair-fed and ad libitum-fed animals. However, animals treated with ethanol for 2, 3, or 4 wk showed decreased hypothalamic and plasma levels of IR-β-EP and decreased splenic NK cell activity. No significant decrease in the number of splenocytes and NK cells or in the percentage of NK cells was seen until after 3 and 4 wk of ethanol treatment. Exposure in vitro of splenic lymphocytes obtained from control animals to various concentrations of β-EP increased NK cell activity. The opiate antagonist naltrexone blocked the β-EP-stimulated effect. The in vitro NK cell response to β-EP was reduced in the splenocytes obtained from animals treated with ethanol for 2 wk, but not in those obtained from animals treated with ethanol for 1 wk as compared with those in control animals. Additionally, β-EP administration into the paraventricular nucleus of the hypothalamus stimulated NK cell cytolytic activity, whereas the opiate blocker administration reduced NK cell activity. The NK cell responses to paraventricular nucleus β-EP were reduced in the animals treated with ethanol for 2 wk. These data provide evidence for the first time that ethanol inhibits NK cell cytolytic activity, possibly by reducing β-EP-regulated splenic NK cell function.

Background Natural killer (NK) cell dysfunction is associated with hyperresponse of corticotropin releasing hormone (CRH) to immune challenge and with a loss of β-endorphin (BEP) neurons in fetal alcohol exposed animals. Recently, we established a method to differentiate neural stem cells into BEP neurons using cyclic adenosine monophosphate (cAMP)-elevating agents in cultures. Hence, we determined whether in vitro differentiated BEP neurons could be used for reversing the compromised stress response and immune function in fetal alcohol exposed rats. Methods To determine the effect of BEP neuron transplants on NK cell function, we implanted in vitro differentiated BEP neurons into the paraventricular nucleus of pubertal and adult male rats exposed to ethanol or control in utero. The functionality of transplanted BEP neurons was determined by measuring proopiomelanocortin (POMC) gene expression in these cells and their effects on CRH gene expression under basal and after lipopolysaccaride (LPS) challenge. In addition, the effectiveness of BEP neurons in activating NK cell functions is determined by measuring NK cell cytolytic activity and interferon-γ (IFN-γ) production in the spleen and in the peripheral blood mononuclear cell (PBMC) following cell transplantation. Results We showed here that when these in vitro differentiated BEP neurons were transplanted into the hypothalamus, they maintain biological functions by producing POMC and reducing the CRH neuronal response to the LPS challenge. BEP neuronal transplants significantly increased NK cell cytolytic activity in the spleen and in the PBMC and increased plasma levels of IFN-γ in control and fetal alcohol exposed rats. Conclusions These data further establish the BEP neuronal regulatory role in the control of CRH and NK cell cytolytic function and identify a possible novel therapy to treat stress hyper-response and immune deficiency in fetal alcohol exposed subjects.

Recent studies show that alterations in the body’s biological rhythms can lead to serious pathologies, including cancer. Acute and chronic ethanol consumption impairs the immune system by causing specific defects in the cellular components of the innate immune response and by creating increased risk and susceptibility to infections and cancer. NK cells are critical for immune surveillance against infected and malignant cells. To assess whether NK cell function follows a circadian trend and to determine ethanol effects on this rhythm, we measured, over a 24-h period, mRNA and protein levels of granzyme B, perforin, and the cytokine IFN-γ, as well as NK cell activity, in the splenocytes of ad libitum-fed, pair-fed, and ethanol-fed Sprague Dawley male rats. Circadian rhythms were found in mRNA and protein levels of granzyme B, perforin, and IFN-γ. A circadian pattern was also detected in NK cell cytolytic activity. Our data further demonstrated how chronic ethanol suppressed NK cell activity by directly disrupting the circadian rhythms of granzyme B, perforin, and IFN-γ. These findings identify the circadian functions of splenic NK cells and show the vulnerability of these rhythms to chronic ethanol.

Background-Microglia are the major inflammatory cells in the central nervous system and play a role in brain injuries as well as brain diseases. In this study, we determined the role of microglia in ethanol's apoptotic action on neuronal cells obtained from the mediobasal hypothalamus and maintained in primary cultures. We also tested the effect of cAMP, a signaling molecule critically involved in hypothalamic neuronal survival, on microglia-mediated ethanol's neurotoxic action.

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Neurobehavioral stress has been shown to promote tumor growth and progression as well as dampen the immune system. In this study, we investigated whether inhibiting stress hormone production could inhibit the development of mammary carcinoma and metastasis in a rat model of breast carcinogenesis. To enhance β-endorphin (BEP), the endogenous opioid polypeptide that boosts immune activity and decreases stress, we generated - BEP neurons by in vitro differentiation from fetal neuronal stem cells and transplanted them into the hypothalami of rats subjected to breast carcinogenesis. BEP transplanted rats displayed a reduction in mammary tumor incidence, growth, malignancy rate, and metastasis compared to cortical cells transplanted rats. BEP neuron transplants also reduced inflammation and epithelial to mesenchymal transition (EMT) in the tumor tissues. In addition, BEP neuron transplants increased peripheral natural killer (NK) cell and macrophage activities, elevated plasma levels of anti-inflammatory cytokines and reduced plasma levels of inflammatory cytokines. Anti-metastatic effects along with stimulation of NK cells and macrophages could be reversed by treatment with the opiate antagonist naloxone, the β-receptor agonist metaproterenol, or the nicotine acetylcholine receptor antagonist methyllycaconitine. Together, our findings establish a protective role for BEP against the growth and metastasis of mammary tumor cells by altering autonomic nervous system activities that enhance innate immune function.

The neurotransmitter dopamine is known to inhibit prolactin secretion and the proliferation of lactotropes in the pituitary gland. In this study, we determined whether dopamine and TGFbeta1 interact to regulate lactotropic cell proliferation. We found that dopamine and the dopamine agonist bromocriptine stimulated TGFbeta1 secretion and TGFbeta1 mRNA expression but inhibited lactotropic cell proliferation both in vivo and in vitro. The dopamine's inhibitory action on lactotropic cell proliferation was blocked by a TGFbeta1-neutralizing antibody. We also found that PR1 cells, which express low amounts of the dopamine D2 receptor, demonstrated reduced expression of TGFbeta1 type II receptor and TGFbeta1 mRNA levels and had undetectable levels of TGFbeta1 protein. These cells showed a reduced TGFbeta1 growth-inhibitory response. Constitutive expression of the D2 receptor short isoform, but not the D2 receptor long isoform, induced TGFbeta1 and TGFbeta1 type II receptor gene expression and recovered dopamine- and TGFbeta1-induced growth inhibition in PR1 cells. The constitutive expression of D2 receptor short isoform also reduced the tumor cell growth rate. These data suggest that a TGFbeta1 system may mediate, in part, the growth-inhibitory action of dopamine on lactotropes.