Induction of astrocyte differentiation by propentofylline increases glutamate transporter expression in vitro: heterogeneity of the quiescent phenotype.
Reactive astrocytes display decreased glutamate transporters, such as GLT-1, and as a result synaptic glutamate clearance is impaired. In addition, these activated astrocytes are immunocompetent and release algesic mediators that can sensitize neurons in the spinal cord. Currently, we evaluated the effect of propentofylline (PPF), an experimental antiallodynic agent, on the phenotype and glutamate transporter expression of astrocytes. Primary astrocyte cultures, which represent an activated phenotype with a polygonal morphology and low GLT-1 expression, were treated for 3 or 7 days with 10, 100, or 1,000 microM PPF or dibutyryl-cAMP (db-cAMP), a known inducer of GLT-1 expression. PPF dose-dependently induced astrocytes to display a mature phenotype, with elongated processes and a stellate shape, as well as increased GLT-1 and GLAST immunoreactivity, similar to that seen with db-cAMP. Real time RT-PCR and Western blot analysis clearly demonstrated that PPF caused a potent dose-dependent induction of GLT-1 and GLAST mRNA and protein in these astrocytes. Importantly, the observed increase in glutamate transporters was found to have a functional effect, with significantly enhanced glutamate uptake in astrocytes treated with 100 or 1,000 microM PPF that was sensitive to dihydrokainate inhibition, suggesting it is GLT-1 mediated. Finally, the effect of PPF on lipopolysaccharide-induced chemokine release was investigated. Interestingly, PPF was able to dampen both MCP-1 (CCL2) and MIP-2 (CXCL2) release from astrocytes while db-cAMP significantly enhanced this chemokine expression. These findings suggest that PPF is capable of differentiating astrocytes to a homeostatic, mature phenotype, competent for glutamate clearance and distinct from that induced by db-cAMP.
Tawfik
Vivianne L
VL
Department of Pharmacology, Dartmouth Medical School, Hanover, New Hampshire 03756, USA.
Lacroix-Fralish
Michael L
ML
Bercury
Kathryn K
KK
Nutile-McMenemy
Nancy
N
Harris
Brent T
BT
Deleo
Joyce A
JA
D027322:Amino Acid Transport System X-AG / Q000235:genetics*; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000166:cytology* / Q000187:drug effects; D001483:Base Sequence; D001692:Biological Transport / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D018698:Glutamic Acid / Q000378:metabolism; D009457:Neuroglia / Q000166:cytology / Q000187:drug effects; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010641:Phenotype; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012964:Sodium / Q000494:pharmacology; D014970:Xanthines / Q000494:pharmacology*
D027322:Amino Acid Transport System X-AG; D017931:DNA Primers; D018696:Neuroprotective Agents; D014970:Xanthines; D018698:Glutamic Acid; C032114:propentofylline; D012964:Sodium