Partial characterization of the neurotrophic factor for maintenance of acetylcholinesterase and butyrylcholinesterase in the preganglionically denervated superior cervical ganglion of the cat in vivo.

Abstract

In continuation of previous reports, it was found that the neurotrophic factor (NF) of the central nervous system of the cat for the maintenance of acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7; AcChoEase) in the denervated cat superior cervical ganglion is a heat-stable compound of low molecular weight (<1,000) and that it is probably a peptide. Acetylcholine and nerve growth factor were eliminated as the NF; cyclic AMP produced an effect similar to that of the NF. The NF is probably not present in significant amounts in liver or skeletal muscle; it appears to be present in small intestine. It does not modify the AcChoEase content of the nondenervated cat superior cervical ganglion. Possible mechanisms of action of the NF are discussed.We have shown that arterial infusion for 24 hr of an extract of the brain, spinal cord, and sciatic nerves of cats prevents almost completely the fall in the acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7; AcChoEase) and butyrylcholinesterase (acylcholine acylhydrolase, EC 3.1.1.8; BtChoEase) contents of the cat superior cervical ganglion (SCG) that otherwise results 48 hr after preganglionic denervation (1). It was shown subsequently that ligation of the external carotid (EC) and lingual (L) arteries is necessary to obtain the full effect of the extract and to counteract partially a similar but lesser effect produced by continual anesthesia with sodium pentobarbital; although the effect was demonstrable when the infusion time was reduced to 6 hr, it was much less marked and consistent than with 24-hr infusion (2). The compound(s) responsible for the preservation of ganglionic AcChoEase and BtChoEase was designated a neurotrophic factor (NF).The present report describes studies addressed to characterization of the NF by 24-hr infusions of authentic agents and of dialyzed and enzyme-treated preparations of the central nervous system (CNS)/sciatic nerve extract. METHODSPreparation of the infusion extract (extract of the brain, spinal cord, and sciatic nerves of cats), anesthetic and surgical procedures, and methods for homogenization of ganglia, for determination of ganglionic AcChoEase, BtChoEase, and protein contents, and for calculation of statistical significance of mean differences were identical with those reported previously (1). Under sodium pentobarbital anesthesia (35 mg/kg of body weight i.p.), 1 cm was resected from both cervical sympathetic trunks; the wound was sutured, and Combiotic (penicillin and dihydrostreptomycin, Pfizer; 0.5 ml) was given i.m. The following day cats were anesthetized as before and treated with atropine (1.0 mg/kg i.p.); artificial respiration was administered through a tracheal catheter attached to a Palmer pump, and a slow i.v. infusion of 5% glucose/0.45% NaCl was started. Heparin at 50 units/kg was given i.v. just prior to bilateral ligation of the EC and L arteries and was repeated every 6 hr. Infusion of the solution under test was begun -24 hr after denervation and continued until the time of sac...

L-glutamic acid, a neurotrophic factor for maintenance of acetylcholinesterase and butyrylcholinesterase in the preganglionically denervated superior cervical ganglion of the cat.

“…Values (Tables 3 and 4 in ref. 5) were considerably higher than those found here or previously (9) with authentic compounds. The present findings are also inconsistent with the earlier conclusion that the endogenous neurotrophic factor is probably a peptide (5).…”

“…Although the preceding and present studies have led to the demonstration that glutamic acid functions as a neurotrophic factor in vivo, they do not of course implicate it as the major compound that is responsible for this effect in dialysates of extracts of the central nervous system (5), or physiologically in the maintenance of postsynaptic AcChoEase and BtChoEase in the normal cat SCG (25). Values (Tables 3 and 4 in ref.…”

“…The neurotrophic factor responsible for this effect was found to be a heat-stable compound of low molecular weight (Mr, <1000) and probably a peptide (5). It was postulated (6) that the neurotrophic factor might act by regulating the conversion of the G1 to the G4 and A12 molecular aggregates of the enzymes (7).…”

Effects of glycyl-L-glutamine in vitro on the molecular forms of acetylcholinesterase in the preganglionically denervated superior cervical ganglion of the cat.

“…In a recent series of experiments (1,(3)(4)(5)(6) it was shown that the 24-hr intracarotid infusion of a dialyzed aqueous extract of cat brain, spinal cord, and sciatic nerves, or of glycyl-L-glutamine (Gly-Gln), or of some closely related compounds, opposes the decrease of both enzymes. Gly-Gln was tested after the demonstration by Haynes and Smith (7) that this compound increases the formation of the G4 (globular tetramer) and A12 (collagen-tailed, asymmetric dodecamer) forms of AcChoEase (8) in cultured preparations of embryonic rat and chicken skeletal muscle.…”

“…The activities of lactate dehydrogenase (LDH) and enolase were determined as indicated previously (9). An aqueous extract of cat central nervous system, which was used in a few experiments, was prepared and dialyzed exactly as described previously (1,3) and was stored at -70°C.…”