GBM (glioblastoma multiforme) is a highly aggressive brain tumour with very poor prognosis despite multi-modalities of treatment. Furthermore, recent failure of targeted therapy for these tumours highlights the need of appropriate rodent models for preclinical studies. In this review, we highlight the most commonly used rodent models (U251, U86, GL261, C6, 9L and CNS-1) with a focus on the pathological and genetic similarities to the human disease. We end with a comprehensive review of the CNS-1 rodent model.

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Antinociceptive tolerance to opioids is a well-described phenomenon, which severely limits the clinical efficacy of opioids for the treatment of chronic pain syndromes. The mechanisms that drive antinociceptive tolerance, however, are less well understood. We have previously shown that glia have a central role in the development of morphine tolerance and that administration of a glial modulating agent attenuated tolerance formation. Recently, we have demonstrated that morphine enhances microglial Iba1 expression and P2X 4 receptor-mediated microglial migration via direct μ opioid receptor signaling in in vitro microglial cultures. We hypothesize that P2X 4 receptors drive morphine tolerance and modulate morphine-induced spinal glial reactivity. Additionally, we hypothesize that perivascular microglia play a role in morphine tolerance and that P2X 4 receptor expression regulates perivascular microglia ED2 expression. To test these hypotheses, rats were implanted with osmotic minipumps releasing morphine or saline subcutaneously for seven days. Beginning three days prior to morphine treatment, P2X 4 receptor antisense oligonucleotide (asODN) was injected intrathecally daily, to selectively inhibit P2X 4 receptor expression. P2X 4 receptor asODN treatment inhibited morphine-induced P2X 4 receptor expression and blocked antinociceptive tolerance to systemically administered morphine. P2X 4 receptor asODN treatment also attenuated the morphine-dependent increase of spinal ionized calcium binding protein (Iba1), glial fibrillary acidic protein (GFAP) and μ opioid receptor protein expression. Chronic morphine also decreased perivascular microglial ED2 expression, which was reversed by P2X 4 receptor asODN. Together, these data suggest that modulation of P2X 4 receptor expression on microglia and perivascular microglia may prove an attractive target for adjuvant therapy to attenuate opioidinduced antinociceptive tolerance.

We recently demonstrated a contributing role of spinal cord infiltrating CD41 T lymphocytes in the maintenance of mechanical hypersensitivity in a rodent model of neuropathic pain, spinal nerve L5 transection (L5Tx). It has been demonstrated that microglia play a role in the etiology of pain states. We hypothesized that infiltrating CD4 1 T lymphocytes communicate with microglia via a CD40-CD154 interaction. Here, we investigated the role of CD40 in the development of mechanical hypersensitivity post-L5Tx. CD40 KO mice displayed significantly decreased mechanical sensitivity compared with WT mice starting from day 5 post-L5Tx. Using bone marrow chimeric mice, we further identified a pro-nociceptive role of CNS microglial CD40 rather than the peripheral leukocytic CD40. Flow cytometric analysis determined a significant increase of CD40 1 microglia in the ipsilateral side of lumbar spinal cord post-L5Tx. Further, spinal cord proinflammatory cytokine profiling demonstrated an induction of IL-6 in both WT and CD40 KO mice post-L5Tx prior to the increase of microglial CD40 expression, indicating a CD40-independent induction of IL-6 following L5Tx. These data establish a novel role of microglial CD40 in the maintenance of nerve injury-induced behavioral hypersensitivity, a behavioral sign of neuropathic pain.Key words: CD40 knockout mice . Bone marrow chimeras . Microglia . IL-6 IntroductionChronic pain is a serious health problem. According to a report released by the Centers for Disease Control and Prevention (CDC) in 2006, one in ten adults in the US suffers from pain that lasts 1 year or more. Neuropathic pain, defined as pain initiated or caused by a primary lesion or dysfunction in the nervous system [1], is one of the most devastating kinds of chronic pain. The concept that microglial activation is crucial in the development of neuropathic pain has been extensively studied and reviewed [2,3]. A phenotype of activated microglia, represented by elevated CD11b, TLR4, B7.2, and MHC II surface expression, has been detected during neuropathic pain development [3,4]. However, despite the emerging findings of a microglial involvement in neuropathic pain, the role of a microglial CD40 signaling pathway has not been investigated in neuropathic pain. The interaction between CD40, a 48 kD cell surface receptor primarily expressed by APC and CD40 ligand (CD154), a 34-39 kD surface protein primarily expressed by activated CD4 1 T lymphocytes, has long been known to play critical roles in both humoral and cell-mediated immune responses [5,6]. Increasingly, CD40-CD154 ligation has been linked to the pathogenesis of various CNS diseases. Elevated CD40 expression was observed during CNS diseases, such as multiple sclerosis, Alzheimer's disease, 3562amyotrophic lateral sclerosis, and HIV-1 encephalitis, both in human patients and in animal models of these CNS diseases [7][8][9][10][11]. Blocking the CD40-CD154 pathway in the CNS resulted in reduced clinical manifestations of multiple sclerosis and Alzheimer's disease in the res...

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain cancer, with a median survival of less than 2 years after diagnosis. The tumor microenvironment plays a critical role in tumor invasion and progression. Microglia and infiltrating macrophages are the most abundant immune cells in the tumor. In the present study, we demonstrate that systemic propentofylline (PPF), an atypical methylxanthine with central nervous system (CNS) glial modulating and anti-inflammatory actions, significantly decreased tumor growth in a CNS-1 rat model of GBM by targeting microglia and not tumor cells. Rats received tumor injections of 1 × 10(5) CNS-1 cells in the right striatum with daily intraperitonial injections of PPF (50 mg/kg) or saline beginning the day of tumor injection. PPF did not cause apoptosis or decrease proliferation of CNS-1 tumor cells. Furthermore, we demonstrate, using in vitro methods, that PPF decreased microglial migration toward CNS-1 tumor cells and decreased MMP-9 expression. The effects of PPF were shown to be specific to microglia and not peripheral macrophages. These results support a differential functional role of resident microglia and infiltrating macrophages in the brain tumor environment. Our data highlight microglia as a crucial target for future therapeutic development and present PPF as a possible drug for treatment of human GBM.

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain cancer, with a median survival of less than 2 years after diagnosis with current available therapies. The tumor microenvironment serves a critical role in tumor invasion and progression, with microglia as a critical player. Our laboratory has previously demonstrated that propentofylline, an atypical methylxanthine with central nervous system glial modulating and anti-inflammatory actions, significantly decreases tumor growth in a GBM rodent model by preferentially targeting microglia. In the present study, we used the CNS-1 rat glioma model to elucidate the mechanisms of propentofylline. Here we demonstrate that propentofylline targets TROY, a novel signaling molecule up-regulated in infiltrating microglia, and not macrophages, in response to CNS-1 cells. We identify Pyk2, Rac1 and pJNK as the downstream signaling molecules of TROY through western blot analysis and siRNA transfection. We demonstrate that inhibition of TROY expression in microglia by siRNA transfection significantly inhibits microglial migration towards CNS-1 cells similar to 10 µM propentofylline treatment. These results identify TROY as a novel molecule expressed in microglia, involved in their migration and targeted by propentofylline. Furthermore, these results describe a signaling molecule that is differentially expressed between microglia and macrophages in the tumor microenvironment.

Glioblastoma multiforme is one of the most common and aggressive primary brain tumors in adults. High glutamate levels are thought to contribute to glioma growth. While research has focused on understanding glutamate signaling in glioma cells, little is known about the role of glutamate between glioma and astrocyte interactions. To study the relationship between astrocytes and tumor cells, the CNS-1 rodent glioma cell line was used. We hypothesized increased glutamate uptake by astrocytes would negatively affect CNS-1 cell growth. Primary rodent astrocytes and CNS-1 cells were co-cultured for 7 days in a Boyden chamber in the presence of 5 mM glutamate. Cells were treated with propentofylline, an atypical synthetic methylxanthine known to increase glutamate transporter expression in astrocytes. Our results indicate astrocytes can increase glutamate uptake through the GLT-1 transporter, leading to less glutamate available for CNS-1 cells, ultimately resulting in increased CNS-1 cell apoptosis. These data suggest that astrocytes in the tumor microenvironment can be targeted by the drug, propentofylline.

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