It has been shown that ethanol exposure can activate astrocytes and microglia resulting in the production of neuroimmune factors, including the chemokine CCL2. The role of these neuroimmune factors in the effects of ethanol on the central nervous system has yet to be elucidated. To address this question, we investigated the effects of ethanol on synaptic transmission and plasticity in the hippocampus from mice that express elevated levels of CCL2 in the brain and their non-transgenic littermate controls. The brains of the transgenic mice simulate one aspect of the alcoholic brain, chronically increased levels of CCL2. We used extracellular field potential recordings in acutely isolated hippocampal slices to identify neuroadaptive changes produced by elevated levels of CCL2 and how these neuroadaptive changes affect the actions of acute ethanol. Results showed that synaptic transmission and the effects of ethanol on synaptic transmission were similar in the CCL2-transgenic and nontransgenic hippocampus. However, long-term potentiation (LTP), a cellular mechanism thought to underlie learning and memory, in the CCL2-transgenic hippocampus was resistant to the ethanol-induced depression of LTP observed in the non-transgenic hippocampus. Consistent with these results, ethanol pretreatment significantly impaired cued and contextual fear conditioning in non-transgenic mice, but had no effect in CCL2-transgenic mice. These data show that chronically elevated levels of CCL2 in the hippocampus produce neuroadaptive changes that block the depressing effects of ethanol on hippocampal synaptic plasticity and support the hypothesis that CCL2 may provide a neuroprotective effect against the devastating actions of ethanol on hippocampal function.

Emerging research provides strong evidence that activation of CNS glial cells occurs in neurological diseases and brain injury and results in elevated production of neuroimmune factors. These factors can contribute to pathophysiological processes that lead to altered CNS function. Recently, studies have also shown that both acute and chronic alcohol consumption can produce activation of CNS glial cells and the production of neuroimmune factors, particularly the chemokine CCL2. The consequences of alcohol-induced increases in CCL2 levels in the CNS have yet to be fully elucidated. Our studies focus on the hypothesis that increased levels of CCL2 in the CNS produce neuroadaptive changes that modify the actions of alcohol on the CNS. We utilized behavioral testing in transgenic mice that express elevated levels of CCL2 to test this hypothesis. The increased level of CCL2 in the transgenic mice involves increased astrocyte expression. Transgenic mice and their non-transgenic littermate controls were subjected to one of two alcohol exposure paradigms, a two-bottle choice alcohol drinking procedure that does not produce alcohol dependence or a chronic intermittent alcohol procedure that produces alcohol dependence. Several behavioral tests were carried out including the Barnes Maze, Y-Maze, cued and contextual conditioned fear test, light-dark transfer, and forced swim test. Comparisons between alcohol naïve, non-dependent, and alcohol dependent CCL2 transgenic and non-tg mice show that elevated levels of CCL2 in the CNS interact with alcohol in tests for alcohol drinking, spatial learning and associative learning.

In the early neonatal period activation of GABA B receptors attenuates calcium current through Ntype calcium channels while enhancing current through L-type calcium channels in rat hippocampal neurons. The attenuation of N-type calcium current has been previously demonstrated to occur through direct interactions of the βγ subunits of G i/o G-proteins, but the signal transduction pathway for the enhancement of L-type calcium channels in mammalian neurons remains unknown. In the present study, calcium currents were elicited in acute cultures from postnatal day 6-8 rat hippocampi in the presence of various modulators of protein kinase A (PKA) and protein kinase C (PKC) pathways. Overnight treatment with an inhibitor of G i/o (pertussis toxin, 200 ng/ml) abolished the attenuation of calcium current by the GABA B agonist, baclofen (10 μM) with no effect on the enhancement of calcium current. These data indicate that while the attenuation of N-type calcium current is mediated by the G i/o subtype of G-protein, the enhancement of L-type calcium current requires activation of a different G-protein. The enhancement of the sustained component of calcium current by baclofen was blocked by PKC inhibitors, GF-109203X (500 nM), chelerythrine chloride (5 μM), and PKC fragment 19-36 (2 μM) and mimicked by the PKC activator phorbol-12-myristate-13-acetate (1 μM). The enhancement of the sustained component of calcium current was blocked by PKA inhibitors H-89 (1 μM) and PKA fragment 6-22 (500 nM) but not Rp-cAMPS (30 μM) and it was not mimicked by the PKA activator, 8-Br-cAMP (500 μM -1 mM). The data suggest that activation of PKC alone is sufficient to enhance L-type calcium current but that PKA may also be involved in the GABA B receptor mediated effect. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access Author ManuscriptNeuroscience. Author manuscript; available in PMC 2012 April 14. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript L-type calcium channels play a large role in the function of both cardiac and skeletal muscle. Thus, many investigators have concentrated on studying the regulation of L-type calcium channels in muscle tissue, particularly cardiac muscle (for review see Catterall, 2000). Most of these physiological studies have categorized current as L-type without attributing it to one of the four specific isoforms of L-type calcium channels that have been sequenced (Catterall, et al., 2005). Since not all isoforms of L-type channels are expressed in all tissues, the tissue type often gives a good indication of the isoform res...

Emerging research has identified that neuroimmune factors are produced by cells of the central nervous system (CNS) and play critical roles as regulators of CNS function, directors of neurodevelopment and responders to pathological processes. A wide range of neuroimmune factors are produced by CNS cells, primarily the glial cells, but the role of specific neuroimmune factors and their glial cell sources in CNS biology and pathology have yet to be fully elucidated. We have used transgenic mice that express elevated levels of a specific neuroimmune factor, the cytokine IL-6 or the chemokine CCL2, through genetic modification of astrocyte expression to identify targets of astrocyte produced IL-6 or CCL2 at the protein level. We found that in non-transgenic mice constitutive expression of IL-6 and CCL2 occurs in the two CNS regions studied, the hippocampus and cerebellum, as measured by ELISA. In the CCL2 transgenic mice elevated levels of CCL2 were evident in the hippocampus and cerebellum, whereas in the IL-6 transgenic mice, elevated levels of IL-6 were only evident in the cerebellum. Western blot analysis of the cellular and synaptic proteins in the hippocampus and cerebellum of the transgenic mice showed that the elevated levels of CCL2 or IL-6 resulted in alterations in the levels of specific proteins and that these actions differed for the two neuroimmune factors and for the two brain regions. These results are consistent with cell specific profiles of action for IL-6 and CCL2, actions that may be an important aspect of their respective roles in CNS physiology and pathophysiology.

During the early postnatal period, GABA(B) receptor activation facilitates L-type calcium current in rat hippocampus. One developmental process that L-type current may regulate is the change in expression of the K(+)Cl(-) co-transporter (KCC2) and N(+)K(+)2Cl(-) co-transporter (NKCC1), which are involved in the maturation of the GABAergic system. The present study investigated the connection between L-type current, GABA(B) receptors, and expression of chloride transporters during development. The facilitation of L-type current by GABA(B) receptors is more prominent in the second week of development, with the highest percentage of cells exhibiting facilitation in cultures isolated from 7 day old rats (37.5%). The protein levels of KCC2 and NKCC1 were investigated to determine the developmental timecourse of expression as well as expression following treatment with an L-type channel antagonist and a GABA(B) receptor agonist. The time course of both chloride transporters in culture mimics that seen in hippocampal tissue isolated from various ages. KCC2 levels increased drastically in the first two postnatal weeks while NKCC1 remained relatively stable, suggesting that the ratio of the chloride transporters is important in mediating the developmental change in chloride reversal potential. Treatment of cultures with the L-type antagonist nimodipine did not affect protein levels of NKCC1, but significantly decreased the upregulation of KCC2 during the first postnatal week. In addition, calcium current facilitation occurs slightly before the large increase in KCC2 expression. These results suggest that the expression of KCC2 is regulated by calcium influx through L-type channels in the early postnatal period in hippocampal neurons.