Effect of nerve growth factor and thyrotropin releasing hormone on cholinergic neurones in developing rat brain reaggregate cultures lesioned with ethylcholine mustard aziridinium.
Foetal rat whole brain reaggregate cultures were prepared in a serum-supplemented (S+) or serum-free medium (S-). Ethylcholine mustard aziridinium (ECMA) was added to the cultures at 9 days in vitro (DIV) at concentrations of 12.5, 25 or 50 microM. Choline acetyltransferase (ChAT) activity was measured at +2, +48 and +96 hr following treatment. In certain experiments the neurotrophic factors, thyrotropin releasing hormone (TRH: 50 micrograms/ml, daily from 9 DIV) or nerve growth factor (NGF: 7S subunit, 5 ng/ml, 0 and +48 hr following ECMA) were added during ECMA treatment. In both types of reaggregate cultured in S+ and S- media there was a 40-80% loss of ChAT activity following ECMA exposure (final concentration = 12.5 microM), presumed to reflect cholinergic cell loss. In both S+ and S- brain reaggregates NGF produced increased ChAT activity with more marked effects in S+ (45-55% increase, +48-96 hr) than in S- medium (20-25% increase, 2-96 hr). No effect on cholinergic muscarinic receptors (specific 3H-QNB binding) was evident after treatment with NGF. TRH had no effect on ChAT activity in the S+ cultures but produced small increases in the S- culture condition (approx 20%, +2-48 hr). Despite a residual "ECMA-resistant" pool of ChAT in the cultures, neither neurotrophic agent was found to cause a reversal of the lesion. In conclusion, the cholinotoxin ECMA appears to produce a cholinergic deficit in both developing S+ and S- reaggregates. This was not reversible by NGF or TRH at the concentrations and under the conditions tested. NGF had marked effects on ChAT activity without affecting muscarinic receptors in untreated developing brain reaggregates cultured in an S+ medium.
Atterwill
C K
CK
Department of Toxicology, Smith Kline & French Research Ltd, Herts, U.K.
Collins
P
P
Meakin
J
J
Pillar
A M
AM
Prince
A K
AK
D000818:Animals; D001388:Aziridines / Q000633:toxicity*; D001389:Azirines / Q000633:toxicity*; D001921:Brain / Q000187:drug effects*; D002478:Cells, Cultured; D002794:Choline / Q000031:analogs & derivatives* / Q000633:toxicity; D002795:Choline O-Acetyltransferase / Q000032:analysis; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009414:Nerve Growth Factors / Q000494:pharmacology*; D010275:Parasympathetic Nervous System / Q000187:drug effects*; D051381:Rats; D011976:Receptors, Muscarinic / Q000187:drug effects; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*
D001388:Aziridines; D001389:Azirines; D009414:Nerve Growth Factors; D011976:Receptors, Muscarinic; D013973:Thyrotropin-Releasing Hormone; C044894:ethylcholine aziridinium; D007770:L-Lactate Dehydrogenase; D002795:Choline O-Acetyltransferase; D002794:Choline