In vitro glial responses to nerve growth factor

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Abstract
Nerve growth factor (NGF) stimulates expression of the low affinity neurotrophin receptor p75NGFR mRNA in primary cultures of neonatal rat cortical type I astrocytes. Nerve growth factor treatment altered glial morphology in glial fibrillary acidic protein positive (GFAP+) cell cultures derived from newborn (P0) and 3-day-old (P3) rat pups. When P0- or P3-derived primary glial cultures were serum-deprived, in the presence of 200 pM NGF for 5 days, the flat polygonal glia present in culture assumed a fibrous morphology, an effect not seen in the untreated serum-deprived controls. The NGF effect on astrocytic morphology was blocked by continuous serum treatment. Nerve growth factor did not stimulate astrocytic proliferation under these culture conditions, as assayed by cell cycle analysis using 3H thymidine autoradiography. P0-derived primary glial cultures expressed the signal transducing neurotrophin receptors p145trkB and p140trka as determined by reverse transcription-polymerase chain reaction (RT-PCR). RT-PCR products were identified by sequencing or restriction enzyme analysis. Astrocytes internalized 125I-NGF at 37°C but not at 4°C, consistent with energy requirements for internalization. Also, internalization of 125I-NGF was abolished by the addition of a 300-1,000-fold excess of unlabeled NGF. Thus, astroglial cells in culture internalize NGF through a specific receptor-mediated process, express trkA and full-length trkB mRNAs at low levels, and respond to exogenous NGF by expressing a fibrous morphology under serum-free culture conditions.