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1.  Differential migration, LPS-induced cytokine, chemokine and NO expression in immortalized BV-2 and HAPI cell lines and primary microglial cultures 
Journal of neurochemistry  2008;107(2):557-569.
Microglial cells are hematopoietically derived monocytes of the CNS and serve important neuromodulatory, neurotrophic and neuroimmune roles. Following insult to the CNS, microglia develop a reactive phenotype, migrate to the site of injury, proliferate, and release a range of proinflammatory, anti-inflammatory and neurotrophic factors. Isolation of primary microglial cell cultures has been an integral step in elucidating the many roles of these cells. In addition to primary microglial cells, several immortalized cell lines have been created to model primary microglia in vitro, including murine derived BV-2 cells and rat derived HAPI cells. Here we compare rat primary microglial, BV-2 and HAPI cells in experiments assessing migration, expression of activation markers and production and release of NO (nitric oxide), cytokines and chemokines. BV-2 and HAPI cells responded similarly to primary microglia in experiments assessing migration, Iba1 expression, and NO release. However, BV-2 and HAPI cells did not model primary microglia in experiments assessing TNFα, IL-1β, IL-6 and MCP-1 expression and release and pERK 44/42 (extracellular receptor kinase) expression following LPS treatment. These results indicate that BV-2 and HAPI cell cultures only partially model primary microglia and that their use should therefore be carefully considered.
PMCID: PMC2581646  PMID: 18717813
BV-2; HAPI; Microglia; Migration; Neuroimmune activation
2.  A Comparison of Spinal Iba1 and GFAP expression in Rodent Models of Acute and Chronic Pain 
Brain research  2008;1219:116-126.
The treatment of acute and chronic pain is still deficient. The modulation of glial cells may provide novel targets to treat pain. We hypothesize that astrocytes and microglia participate in the initiation and maintenance of both, acute surgical and chronic neuropathic pain. Rats underwent paw incision, L5 nerve exposure or L5 nerve transection surgery. Behavioral mechanical allodynia was assessed using von Frey filaments. Immunohistochemistry was performed using anti-ionized calcium binding adaptor protein, Iba-1 (microglia), and anti-Glial Fibrillary Acidic Protein, GFAP (astrocytes) on day 1, 4 and 7 after surgery. Following paw incision and at spinal L5 segment GFAP expression was increased in laminae I-II and Iba1 in deep laminae on day 1, in the entire dorsal horn on day 4 and dissipate on day 7 after paw incision in parallel with the allodynia. L5 nerve transection induced mechanical allodynia from day 1 to 7 which correlated with Iba-1 increases on day 1, 4 (entire dorsal horn) and day 7 after nerve injury (deep laminae of the dorsal horn) at spinal L5 segment. Conversely, GFAP increased at later time points from day 4 (deep laminae) and on day 7 (entire dorsal horn). Our data demonstrates that astrocytes (GFAP expression) play a role in the initiation of acute pain and the maintenance of chronic pain while Iba-1 increases closely correlated with the early phase of neuropathic pain. Iba1 and GFAP increased rostrally, at L3 segment, after paw incision (day 4) and only Iba1 increased following L5 nerve transection (day 7).
PMCID: PMC2512259  PMID: 18538310
paw incision; neuropathic pain; postoperative pain; spinal cord; allodynia; glia
3.  Propentofylline-Induced Astrocyte Modulation Leads to Alterations in Glial Glutamate Promoter Activation Following Spinal Nerve Transection 
Neuroscience  2008;152(4):1086-1092.
We have previously shown that the atypical methylxanthine, propentofylline, reduces mechanical allodynia after peripheral nerve transection in a rodent model of neuropathy. In the present study, we sought to determine whether propentofylline-induced glial modulation alters spinal glutamate transporters, GLT-1 and GLAST in vivo, which may contribute to reduced behavioral hypersensitivity after nerve injury. In order to specifically examine the expression of the spinal glutamate transporters, a novel line of double transgenic GLT-1-eGFP/GLAST-DsRed promoter mice was used. Adult mice received propentofylline (10 mg/kg) or saline via intraperitoneal injection starting 1-hour prior to L5-spinal nerve transection and then daily for 12 days. Mice receiving saline exhibited punctate expression of both eGFP (GLT-1 promoter activation) and DsRed (GLAST promoter activation) in the dorsal horn of the spinal cord, which was decreased ipsilateral to nerve injury on day 12. Propentofylline administration reinstated promoter activation on the injured side as evidenced by an equal number of eGFP (GLT-1) and DsRed (GLAST) puncta in both dorsal horns. As demonstrated in previous studies, propentofylline induced a concomitant reversal of L5 spinal nerve transection-induced expression of Glial Fibrillary Acidic Protein (GFAP). The ability of propentofylline to alter glial glutamate transporters highlights the importance of controlling aberrant glial activation in neuropathic pain and suggests one possible mechanism for the anti-allodynic action of this drug.
PMCID: PMC2423012  PMID: 18358622
Spinal glia; Neuropathic pain; Neuroimmune; Peripheral nerve injury; Mice
4.  Spinal Microglial and Perivascular Cell Cannabinoid Receptor Type 2 Activation Reduces Behavioral Hypersensitivity without Tolerance after Peripheral Nerve Injury 
Anesthesiology  2008;108(4):722-734.
Cannabinoids induce analgesia by acting on cannabinoid receptor (CBR) types 1 and/or 2. However, central nervous system side effects and antinociceptive tolerance from CBR1 limit their clinical use. CBR2 exist on spinal glia and perivascular cells, suggesting an immunoregulatory role of these receptors in the central nervous system. Previously, the authors showed that spinal CBR2 activation reduces paw incision hypersensitivity and glial activation. This study tested whether CBR2 are expressed in glia and whether their activation would induce antinociception, glial inhibition, central side effects, and antinociceptive tolerance in a neuropathic rodent pain model.
Rats underwent L5 spinal nerve transection or sham surgery, and CBR2 expression and cell localization were assessed by immunohistochemistry. Animals received intrathecal injections of CBR agonists and antagonists, and mechanical withdrawal thresholds and behavioral side effects were assessed.
Peripheral nerve transection induced hypersensitivity, increased expression of CR3/CD11b and CBR2, and reduced ED2/CD163 expression in the spinal cord. The CBR2 were localized to microglia and perivascular cells. Intrathecal JWH015 reduced peripheral nerve injury hypersensitivity and CR3/CD11b expression and increased ED2/CD163 expression in a dose-dependent fashion. These effects were prevented by intrathecal administration of the CBR2 antagonist (AM630) but not the CBR1 antagonist (AM281). JWH015 did not cause behavioral side effects. Chronic intrathecal JWH015 treatment did not induce antinociceptive tolerance.
These data indicate that intrathecal CBR2 agonists may provide analgesia by modulating the spinal immune response and microglial function in chronic pain conditions without inducing tolerance and neurologic side effects.
PMCID: PMC2647363  PMID: 18362605
5.  Differential regulation of neuregulin 1 expression by progesterone in astrocytes and neurons 
Neuron glia biology  2006;2(4):227-234.
Glial–neuronal interactions are crucial processes in neuromodulation and synaptic plasticity. The neuregulin 1 family of growth and differentiation factors have been implicated as bidirectional signaling molecules that are involved in mediating some of these interactions. We have shown previously that neuregulin 1 expression is regulated by the gonadal hormones progesterone and 17β-estradiol in the CNS, which might represent a novel, indirect mechanism of the neuromodulatory actions of these gonadal hormones. In the present study, we sought to determine the effects of progesterone and 17β-estradiol on neuregulin 1 expression in rat cortical astrocytes and neurons in vitro. We observed that progesterone increased the expression of neuregulin 1 mRNA and protein in a dose-dependent manner in cultured astrocytes, which was blocked by the progesterone receptor antagonist RU-486. In contrast, 17β-estradiol did not increase either neuregulin 1 mRNA or protein in astrocytes. We observed no effect of either progesterone or 17β-estradiol on neuregulin 1 mRNA and protein in rat cortical neurons in vitro. Finally, we observed that treatment of cortical neurons with recombinant NRG1-β1 caused PSD-95 to localize in puncta similar to that observed following treatment with astrocyte-conditioned medium. These results demonstrate that progesterone regulates neuregulin 1 expression, principally in astrocytes. This might represent a novel mechanism of progesterone-mediated modulation of neurotransmission through the regulation of astrocyte-derived neuregulin 1.
PMCID: PMC2099160  PMID: 18049715
Sex hormones; neuregulin; glial activation; PSD-95; NGR1

Results 1-5 (5)