Astroglial Cell Proliferation Research Articles

Acute and chronic effects of exposure to a 1-mT magnetic field on the cytoskeleton, stress proteins, and proliferation of astroglial cells in culture

This paper reports the effects of exposure to static, sinusoidal (50 Hz), and combined static/sinusoidal magnetic fields on cultured astroglial cells. Confluent primary cultures of astroglial cells were exposed to a 1-mT sinusoidal, static, or combined magnetic field for 1 h. In another experiment, cells were exposed to the combined magnetic field for 1, 2, and 4 h. The hsp25, hsp60, hsp70, actin, and glial fibrillary acidic protein contents of the astroglial cells were determined by immunoblotting 24 h after exposure. No significant differences were seen between control and exposed cells with respect to their contents of these proteins, neither were any changes in cell morphology observed. In a third experiment to determine the effect of a chronic (11-day) exposure to a combined 1-mT static/sinusoidal magnetic field on the proliferation of cultured astroglial cells, no significant differences were seen between control, sham-exposed, or exposed cells. These results suggest that exposure to 1-mT sinusoidal, static, or combined magnetic fields has no significant effects on the stress, cytoskeletal protein levels in, or proliferation of cultured astroglial cells.

Postischemic mild hypothermia reduces neurotransmitter release and astroglial cell proliferation during reperfusion after asphyxial cardiac arrest in rats

The present study investigated whether postischemic mild hypothermia attenuates the ischemia-induced striatal glutamate (GLU) and dopamine (DA) release, as well as astroglial cell proliferation in the brain. Anesthetized rats were exposed to 8 min of asphyxiation, including 5 min of cardiac arrest. The cardiac arrest was reversed to restoration of spontaneous circulation (ROSC), by brief external heart massage and ventilation within a period of 2 min. After the insult and during reperfusion, the extracellular glutamate and dopamine overflow increased to, respectively, 3000% and 5000% compared with the baseline values in the normothermic group and resulted in brain damage, ischemic neurons and gliosis. However, when hypothermia was induced for a period of 60 min after the insult and restoration of spontaneous circulation, the glutamate and dopamine overflows were not significantly different from that in the sham group. Histological analysis of the brain showed that postischemic mild hypothermia reduced brain damage, ischemic neurons, as well as astroglial cell proliferation. Thus, postischemic mild hypothermia reduces the excitotoxic process, brain damage, as well as astroglial cell proliferation during reperfusion. Moreover, these results emphasize the trigger effect of dopamine on the excitotoxic pathway.

Role of phospholipase D signaling in ethanol-induced inhibition of carbachol-stimulated DNA synthesis of 1321N1 astrocytoma cells.

Inhibition of astrocyte proliferation has been suggested to be an important event in the developmental neurotoxicity associated with ethanol. We have previously shown that the acetylcholine analog carbachol induces astroglial cell proliferation through activation of muscarinic M3 receptors, and that ethanol strongly inhibits this effect by inhibiting activation of protein kinase C (PKC) zeta and its down-stream effector 70-kDa ribosomal S6 kinase (p70S6K). In this study, we investigated whether inhibition by ethanol of this signal transduction pathway in 1321N1 human astrocytoma cells may be due, at least in part, to inhibition of the formation of the PKC zeta activator phosphatidic acid (PA), which is formed by hydrolysis of phosphatidylcholine by phospholipase D (PLD). 1-Butanol, which is a substrate for PLD and inhibits PA formation, inhibited carbachol-induced cell proliferation and the underlying intracellular signaling, whereas its analog tert-butanol, which is a poor substrate for PLD, was much less effective. In addition, exogenous PAs were able to increase DNA synthesis and to activate PKC zeta and p70S6K. Furthermore, in carbachol-stimulated cells, ethanol increased the formation of phosphatidylethanol and inhibited the formation of PA. Taken together, these results indicate that PLD activation plays an important role in carbachol-induced astroglial cell proliferation by generating the second messenger PA, which activates PKC zeta. Moreover, the effect of ethanol on carbachol-induced proliferation appears to be mediated, at least in part, by its ability to interact with PLD leading to a decreased synthesis of PA.

Signal transduction mechanisms involved in the antiproliferative effects of ethanol in glial cells

Central nervous system dysfunctions (most notably microencephaly and mental retardation) are among the most significant effects of in utero exposure to ethanol. Ethanol causes alterations of both neuronal and glial cells. In particular, ethanol has been shown to inhibit proliferation of astroglial cells stimulated by certain, but not all mitogens. Here, we review evidence that acetylcholine, by activating the M 3 subtype of muscarinic receptors, increases DNA synthesis in rat and human astroglial cells and that this effect is inhibited by low ethanol concentrations (10–100 mM). Of the several signal transduction pathways activated by these receptors in astrocytes or astrocytoma cells, ethanol appears to target activation of phospholipase D, leading to a decrease in phosphatidic acid, a decreased activation of the atypical protein kinase C ζ and decreased down-stream activation of p70S6 kinase and of nuclear factor-κB. Inhibition of this pathway by ethanol occurs at the same concentrations which effectively inhibit proliferation. Inhibition of astroglial cell proliferation by ethanol may contribute to the microencephaly present in most children diagnosed with the fetal alcohol syndrome.

Growth factor-estradiol interaction on DNA labeling and cytoskeletal protein expression in cultured rat astrocytes

Growth factors are major signaling agents regulating neuron-glia dialogue. Epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), insulin growth factor-I (IGF-I) and insulin (INS) induce neuronal and astroglial cell proliferation and differentiation. This is true also for estrogens that influence astrocytes and exert neuroprotectant activity. In this study interactions between growth factors and estradiol on DNA labeling and glial fibrillary acidic protein (GFAP) and vimentin expression in cultured astrocytes were investigated. DNA labeling was significantly stimulated by bFGF pretreatment followed by 24 h estradiol and EGF or IGF-I or INS added in the last 12 h. Western blotting showed also a modulation of GFAP and vimentin expression in treated astrocytes. This suggests the occurrence of a crucial growth factor-estradiol interaction on DNA labeling and cytoskeletal protein expression during astrocyte proliferation and differentiation in culture.

Glial Fibrillary Acidic Protein and Vimentin Expression Is Regulated by Glucocorticoids and Neurotrophic Factors in Primary Rat Astroglial Cultures

The neurotrophic factors epidermal growth factor (EGF), basic fibroblast growth factor, (bFGF), insulin‐like growth factor I (IGF‐I) and insulin (INS) regulate neural and astroglial cell functions. Glucocorticoids may influence the metabolism of astroglial compartment and are key hormones in neurodegenerative events. This study was designed to assess the interactions between growth factors and dexamethasone (DEX) on cytoskeletal proteins (GFAP and vimentin) expression in 25 days in vitro (DIV) astrocyte cultures. An increase in GFAP and vimentin expression was observed after 12 h pretreatment with bFGF and subsequent treatment for 60 h with DEX. GFAP immunoreactivity was decreased after 24 h progression growth factors (EGF, IGF‐I and INS) addition, when compared to control 36 h DEX and bFGF‐pretreated cultures for the last 12 h. Vimentin immunoreactivity was decreased after 12 h bFGF pretreatment and subsequent 60 h DEX addition in astrocyte cultures compared to 12 h bFGF‐pretreated ones. Pretreatment for 36 h with DEX plus bFGF in the last 12 h and subsequent treatment for 24 h with DMEM (Dulbecco's modified Eagle medium; DMEM) + BSA (bovine serum albumine) (harvesting), or with progression growth factors (EGF, IGF‐I or INS) alone or two of them together, stimulated GFAP expression, compared to untreated controls. Immunochemical analysis of the mitogen‐activated protein kinase ERK2 suggests an involvement of this enzyme in the control of GFAP expression. The above findings support the view of an interactive and complex dialogue between growth factors and glucocorticoids during astroglial cell proliferation and maturation in culture. This may have implications in therapeutic approach of neurologic disorders associated with astrogliosis, including cerebrovascular disease.

Mitogenic effects of phospholipase D and phosphatidic acid in transiently permeabilized astrocytes: effects of ethanol.

Investigations of lipid-mediated signalling pathways are often limited by a lack of methods for the intracellular delivery of lipid messengers. We established a procedure for the transient permeabilization of astrocytes by an oxygen-insensitive mutant of streptolysin-O (SLO) to investigate the participation of the phospholipase D (PLD) signalling pathway in astroglial cell proliferation. Exogenous PLD, when incubated in the presence of SLO, caused an increase in DNA synthesis (measured by thymidine incorporation) which was completely suppressed by ethanol (0.3%, v/v). In parallel experiments, phosphatidic acid also induced a dose-dependent mitogenic response which, however, was not affected by the presence of ethanol. Phosphatidic acid was more effective in this assay than diacylglycerol but its effect was sensitive to the protein kinase inhibitor Ro 31-8220. Our findings provide direct evidence that disruption of the PLD signalling pathway by ethanol is sufficient to suppress astroglial proliferation, an effect that might contribute to the inhibition of brain growth in alcoholic embryopathy.

Nuclear factor κb activation by muscarinic receptors in astroglial cells: effect of ethanol

Activation of muscarinic receptors leads to proliferation of astroglial cells and this effect is inhibited by ethanol. Among the intracellular pathways involved in the mitogenic action of muscarinic agonists, activation of the atypical protein kinase C ζ (PKC ζ) appears to be of most importance, and is also affected by low ethanol concentrations. PKC ζ has been reported to activate nuclear factor κB (NF-κB), a transcription factor that has been shown to play an important role in cell proliferation. The aim of this study was, therefore, to determine whether muscarinic receptors would activate NF-κB in astroglial cells, whether such activation would play a role in the mitogenic action of muscarinic agonists, and whether it would represent a possible target for ethanol. Carbachol activated NF-κB in human 1321N1 astrocytoma cells, as evidenced by translocation of the p65 subunit of NF-κB to the nucleus, phosphorylation and degradation of IκBα in the cytosol, and increase NF-κB binding to DNA. Carbachol also induced translocation of p65 to the nucleus in primary rat astrocytes. Carbachol-induced NF-κB activation was mediated by the M3 subtype of muscarinic receptors and appeared to involve Ca2+ mobilization and activation of PKC ϵ and PKC ζ, but not PI3-kinase and mitogen-activated protein kinase. The NF-κB peptide inhibitor SN50, but not the inactive peptide SN50M, strongly inhibited carbachol-induced astrocytoma cells proliferation and p65 translocation to the nucleus. Increased DNA synthesis was also antagonized by the IκBα kinase inhibitor BAY 11–7082. Ethanol (25–100 mM) inhibited the translocation of p65 and the binding of NF-κB to DNA in both 1321N1 astrocytoma cells and primary rat cortical astrocytes. Together, these results suggest that activation of NF-κB by muscarinic receptors in astroglial cells is important for carbachol-induced DNA synthesis and that ethanol-mediated inhibition of cell proliferation may be due in part to inhibition of NF-κB activation.

Effect of ethanol on protein kinase Cζ and p70S6 kinase activation by carbachol: a possible mechanism for ethanol‐induced inhibition of glial cell proliferation

AbstractThe signal transduction pathways that mediate the mitogenic response of muscarinic acetylcholine receptors in astroglial cells have not been fully elucidated. In this study we investigated the activation of p70S6 kinase (p70S6K) by carbachol in 1321 N1 astroctyoma cells. Carbachol induced a dose‐ and time‐dependent activation of p70S6K, as evidenced by increased phosphorylation at Thr‐389, Thr‐421 and Ser‐424, by increased p70S6K activity, and by a shift in its molecular weight. Activation of p70S6K was mediated by M3 muscarinic acetylcholine receptors (mAChRs) and was inhibited by two phosphatidylinositol‐3‐kinase (PI3‐K) inhibitors, by a pseudosubstrate to protein kinase C (PKC) ζ, and by the p70S6K inhibitor rapamycin. Carbachol‐induced DNA synthesis was strongly inhibited by rapamycin, suggesting that p70S6K activation plays an important role in carbachol‐induced cell proliferation. Ethanol (25–100 mm) has been shown to inhibit carbachol‐induced proliferation of astroglial cells. In the same range of concentrations, ethanol also inhibits carbachol‐induced activation of PKCζ and of p70S6K. On the other hand, inhibition of PI3‐kinase was only observed at higher ethanol concentrations. These results indicate that activation of the PKCζ→ p70S6K pathway by M3 mAChRs may play a role in the increased DNA synthesis and may represent a target for ethanol‐induced inhibition of astroglial cell proliferation.

EFFECT OF GROWTH FACTORS ON DNA LABELING AND CYTOSKELETAL PROTEIN EXPRESSION IN 17-β-ESTRADIOL AND BASIC FIBROBLAST GROWTH FACTOR PRE-TREATED ASTROCYTE CULTURES

Astrocytes react to all noxae which damage neurons. Their reactions include degeneration, hypertrophy, hyperplasia and fibre formation. Growth factors inducing proliferation and differentiation of both neurons and astrocytes in culture play a pivotal role in the dynamic flow of signaling molecules between neurons and astroglia. Estrogens as well influence astroglia and are neuroprotectants. This study has investigated the interactions between growth factors and estrogens on DNA labeling and cytoskeletal protein [glial fibrillary acidic protein (GFAP) and vimentin] expression in 22 DIV astrocyte cultures treated for 24 or 36 h under different experimental conditions.Contemporary addition of 17-β-estradiol (E2) with two or three growth factors for 24 h, significantly stimulated methyl-[3H]thymidine incorporation into DNA from 22 days in vitro (DIV) astrocyte cultures.This effect reached a peak when E2 was co-added with epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and insulin. In astrocyte cultures treated for 36 h with E2 and EGF+insulin or bFGF+insulin added in the last 12 h, DNA labeling was remarkably increased. The parallel cyclin D1 expression positively correlated with ERK2 activation. Western blot analysis for cytoskeletal proteins showed also changes of both GFAP and vimentin expression. The above data suggest the occurrence of a scheduled interaction between “competence” or “progression” growth factors and estrogens on DNA labeling and cytoskeletal protein expression during astroglial cell proliferation and differentiation in culture. A better understanding of the mechanisms of these interactions may contribute to develop strategies for controlling astroglial reaction in cerebrovascular disease including stroke and hypertensive brain damage.

Activation of phosphatidylinositol 3 kinase by muscarinic receptors in astrocytoma cells.

Stimulation of Gq-coupled acetylcholine muscarinic receptors leads to proliferation of astroglial cells, but the signal transduction pathway(s) that mediate this mitogenic response have not been fully elucidated. In this study, we report on the ability of carbachol to stimulate the phosphorylation of Akt/PKB, an important target of phosphatidylinositol 3 kinase (PI3 kinase) in 1321N1 human astrocytoma cells. Carbachol induced a dose-dependent phosphorylation of Ser473 on Akt, peaking after 15 min. This effect was mediated by activation of the M3 subtype of muscarinic receptors and was inhibited by two PI3 kinase inhibitors. Inhibitors of protein kinase C, mitogen-activated protein kinase and p70S6 kinase, had no effect on carbachol-induced Akt phosphorylation. Carbachol-induced DNA synthesis was strongly inhibited by two PI3 kinase inhibitors, wortmannin and LY294002, suggesting that PI3 kinase activation plays an important role in carbachol-induced proliferation 1321N1 astrocytoma cells.

Regulated expression of a dominant negative protein kinase C ε mutant inhibits the proliferation of U-373MG human astrocytoma cells

The tight regulation of protein kinase C (PKC) activity is crucial for maintaining normal cell proliferation. Excessive PKC activity leads to uncontrolled growth and malignant transformation. It has been reported that the activity of PKC is higher in astroglial cell lines than in normal astrocytes. Previously, we demonstrated that PKC epsilon is overexpressed in astroglial cell lines and in samples from primary high-grade astroglial brain tumors. Because there are no PKC epsilon isozyme-specific inhibitors, we chose a genetic approach to confirm that PKC epsilon is a potential target for inhibiting astroglial cell proliferation. We regulated the expression of a dominant negative PKC epsilon mutant (PKC epsilon 1-401 encoding amino acid 1-401) in U-373MG human astrocytoma cells using a tetracycline-regulated expression vector and established stable clones. Induction of expression of the dominant negative PKC epsilon mutant by the addition of doxycycline, a tetracycline derivative, completely blocked proliferation of U-373MG cells in proliferation and clonogenic assays. Although the induction of the dominant negative PKC epsilon mutant did not markedly affect mitogen-induced tyrosine phosphorylation of the mitogen-activated protein (MAP) kinases, it inhibited the induction of c-Fos protein expression by substance P (SP) and fetal bovine serum (FBS). These results clearly show that the expression of dominant negative PKC epsilon leads to the inhibition of U-373MG cellular proliferation demonstrating that this isozyme may be a potential therapeutic target for astroglial brain tumors.

Morphological abnormalities in the brains of estrogen receptor beta knockout mice.

Estrogen receptor beta (ERbeta) is expressed at high levels in both neurons and glial cells of the central nervous system. The development of ERbeta knockout (BERKO) mice has provided a model to study the function of this nuclear receptor in the brain. We have found that the brains of BERKO mice show several morphological abnormalities. There is a regional neuronal hypocellularity in the brain, with a severe neuronal deficit in the somatosensory cortex, especially layers II, III, IV, and V, and a remarkable proliferation of astroglial cells in the limbic system but not in the cortex. These abnormalities are evident as early as 2 mo of age in BERKO mice. As BERKO mice age, the neuronal deficit becomes more pronounced, and, by 2 yr of age, there is degeneration of neuronal cell bodies throughout the brain. This is particularly evident in the substantia nigra. We conclude that ERbeta is necessary for neuronal survival and speculate that this gene could have an important influence on the development of degenerative diseases of the central nervous system, such as Alzheimer's disease and Parkinson's disease, as well as those resulting from trauma and stroke in the brain.

Inhibition of astroglial cell proliferation by alcohols: interference with the protein kinase C-phospholipase D signaling pathway

Ethanol inhibits astroglial cell proliferation, an effect that may contribute to the development of alcoholic embryopathy in humans. In the present study, we investigated inhibitory effects of ethanol and butanol isomers (1-, 2- and t-butanol) on astroglial cell proliferation induced by the strongly mitogenic phorbol ester, 4ß-phorbol-12α,13ß-dibutyrate (PDB). 4ß-Phorbol-12α,13ß-dibutyrate (PDB) induced a 10-fold increase of [3H]thymidine incorporation in cortical astrocytes prepared from newborn rats (EC50: 70 nM) which was blocked by Ro 31-8220, a cell-permeable protein kinase C (PKC) inhibitor. Ethanol blocked PDB-induced astroglial proliferation in a concentration-dependent manner; significant effects were already seen at 0.1% (v/v). Concomitantly, ethanol caused the formation of phosphatidylethanol (PEth) by phospholipase D (PLD) and reduced PLD-mediated formation of phosphatidic acid (PA). The butanols also inhibited the mitogenic action of phorbol ester; the inhibitory potency of the butanols was 1-butanol>2-butanol>t-butanol. The same range of potencies was observed for the inhibitory activity of the butanols towards protein kinase C activity measured in vitro. At 0.3% concentration, 1-butanol potently suppressed the PDB-induced formation of phosphatidic acid while 2- and t-butanol were less active. Taken together, our results suggest that ethanol and 1-butanol exert a specific inhibitory effect on PKC-dependent astroglial cell proliferation by synergistically inhibiting PKC activity and the PLD signaling pathway.

Functions of transforming growth factor-beta isoforms in the nervous system. Cues based on localization and experimental in vitro and in vivo evidence.

This review briefly describes the cellular distribution and documented roles of the transforming growth factor (TGF)-beta isoforms TGF-beta2 and -beta3 in the central and peripheral nervous system. TGF-beta2 and -beta3 are coexpressed in developing radial glial and mature astroglial and Schwann cells, as well as in subpopulations of differentiated neurons, most prominently in cortical, hippocampal, and brainstem/spinal cord motor neurons. In vitro studies have suggested a number of potential, physiologically relevant functions for TGF-betas including regulation of astroglial cell proliferation, expression of adhesion molecules, survival promoting roles for neurons in combination with established neurotrophic factors, and differentiative actions on neurons.

Extracellular S100B protein modulates ERK in astrocyte cultures.

S100B is a calcium binding protein expressed and secreted by astrocytes. Extracellular S100B stimulates the proliferation of astroglial cells and the survival of neurons. Extracellular signal regulated kinases (ERK) are involved in the transduction of proliferating signals in astrocytes. Here we report that S100B significantly increases the activity of ERK in primary cultures of astrocytes, a result which may be related to previous observations of the effect of this protein on glial proliferation. We further confirm that conversion of S100B to its covalent dimer by oxidation of cysteine residues increases its extracellular activity. Although we cannot exclude S100B involvement in other mechanisms of signal transduction, these results suggest that ERK activity in astrocytes is modulated by extracellular S100B.

Ethanol inhibits astroglial cell proliferation by disruption of phospholipase D-mediated signaling.

The activation of phospholipase D (PLD) is a common response to mitogenic stimuli in various cell types. As PLD-mediated signaling is known to be disrupted in the presence of ethanol, we tested whether PLD is involved in the ethanol-induced inhibition of cell proliferation in rat cortical primary astrocytes. Readdition of fetal calf serum (FCS) to serum-deprived astroglial cultures caused a rapid, threefold increase of PLD activity and a strong mitogenic response; both effects were dependent on tyrosine kinases but not on protein kinase C. Ethanol (0.1-2%) suppressed the FCS-induced, PLD-mediated formation of phosphatidic acid (PA) as well as astroglial cell proliferation in a concentration-dependent manner. Moreover, exogenous bacterial PLD increased astroglial proliferation in an ethanol-sensitive manner, whereas exogenous PA or lysophosphatidic acid was less effective. Formation of PA and astroglial proliferation were strongly inhibited by 1-butanol (0.1-1%), a substrate of PLD, but were unaffected by t-butanol, a non-substrate; 2-butanol had intermediate effects. Platelet-derived growth factor and endothelin-1 mimicked the mitogenic effect of FCS; their effects were also inhibited by the butanols in the potency order 1-butanol > 2-butanol > tert-butanol. Our results, in particular, the differential effects of 1-, 2-, and tert-butanol with respect to PA formation and astroglial proliferation, strongly suggest that the antiproliferative effects of ethanol in glial cells are due to the disruption of the PLD signaling pathway. This mechanism may also contribute to the inhibition of astroglial growth and brain development observed in alcoholic embryopathy.

Loss of Neurofibromin in the Leptomeningeal Astroglial Heterotopia of NF-1

Neurofibromin, the protein product of the neurofibromatosis type 1 (NF-1) gene, has important roles in tumor suppression or normal embryogenesis. Cerebellar leptomeningeal astroglial heterotopia (LAH) is a proliferation of heterotopic astroglial cells and fibers in the cerebellar leptomeninges, which is characteristically demonstrated in the NF-1 patients. In this study, neurofibromin expression was investigated in NF-1 and non-NF-1 human tissues, especially in the cerebellum of NF-1 patients. Neurofibromin was found by immunoblotting in the CNS but not in the heart, liver, and kidney. Immunohistochemistry in the normal areas of the brains with NF-1 demonstrated neurofibromin immunoreactivity as did the brains of unaffected controls. Cerebellar LAH showed no neurofibromin immunoreactivity. The results of this study suggest that neurofibromin expression remains unchanged in the nonproliferated region of the CNS of the NF-1 patient but changes occur in the abnormally proliferated region, resulting in cerebellar LAH. Loss of neurofibromin may result in the excessive migration and growth of astrocytes in the early fetal period.

Kami-Untan-To enhances choline acetyltransferase and Nerve growth factor mRNA levels in brain cultured cells

Effects of a Kampo (Japanese herbal) medicine "kami-untan-to (KUT)" on central cholinergic neurons were examined in vitro. When rat embryo basal forebrain cells were cultured in the presence of KUT (200 μg/ml) for three days, a significant increase of choline acetyltransferase (ChAT) activity was observed. When rat embryo astroglial cells were grown in culture in the presence of KUT (12.5-400 μg/ml) for over 24 hours, the amount of NGF secreted in quiescent astrocytes was increased in a dose-dependent manner; this was detected by the ELISA method. However, KUT did not affect the proliferation of astroglial cells. KUT-induced ChAT activity and NGF secretion were completely blocked in the presence of actinomycin D and/or cycloheximide without a loss in cell viability. The treatment with KUT increased levels of ChAT mRNA in basal forebrain cells and NGF mRNA in cerebral cortex cells, respectively, these results were confirmed by using the RT-PCR (reverse transcriptase-polymerase chain reaction) method. These results suggest that KUT-induced ChAT activity and NGF secretion occur by modulating the transcriptional stage of both proteins.

β-adrenergic receptors regulate astrogliosis and cell proliferation in the central nervous system in vivo

Astrocytes express several cell surface receptors including the beta 2 -adrenergic receptor. To explore whether beta-adrenergic receptors (beta-ARs) directly regulate astrogliosis and glial scar formation, we evaluated the effects of beta-AR activation and blockade on astrocyte hypertrophy and cell proliferation in rabbit optic nerves in vivo. Artificial cerebrospinal fluid (CSF), isoproterenol (ISO; a beta-agonist), or propranolol (PROP; a beta-antagonist) were infused via osmotic minipumps into non-injured and crushed optic nerves for 14 days. Changes in nerve cell numbers and astroglial hypertrophy were monitored by ethidium bromide nuclear staining and glial fibrillary acidic protein (GFAP) immunohistochemistry, respectively. In non-injured nerves infused with CSF or PROP, there were no alterations in GFAP-immunoreactivity or cell numbers compared to normal optic nerves; however, in non-injured nerves infused with ISO, there was a significant increase in both GFAP-immunoreactivity and cell number. In crushed optic nerves, there was a significant increase in both GFAP-immunoreactivity and cell number compared to normal nerves, and this increase was not altered by infusion of either CSF or ISO. In contrast, PROP infusion significantly reduced the crush-induced increase in GFAP-immunofluorescence and cell number. These findings suggest that a) beta-AR activation, in the absence of injury, can promote astroglial hypertrophy and cell proliferation; b) after injury, beta-AR activation drives injury-induced astrogliosis and cell proliferation; c) astrocyte beta-ARs are maximally stimulated after neuronal injury; and d) neuronal regeneration may be influenced, both positively and negatively, through the pharmacological manipulation of glial receptors.