Metabolism and mode of action 



Physostigmine is somewhat related in structure to lAA: 



CH3NH CH3 



COO 



C CH2 



C\ /CH2 



N^ H ^N^ 



II 

 CH3 CH3 physostigmine 



and the finding that lAA acted rather similarly as an apparent acetylcholine- 

 esterase inhibitor in sphenic-diaphragm preparations was consequently 

 not completely surprising. The concentration at which this inhibition 

 was observed was relatively high, 5 X 10 ^ M or around 100 p. p.m. 



It has been shown by Koch (cf. review by him, 1954) that uptake of ions 

 by gills of the crab, Eriocheir, is reversibly inhibited by physostigmine. The 

 relatively irreversible choline-esterase inhibitor, tetraethylpyrophosphate 

 (TEPP), similarly caused irreversible inhibition in this particular tissue, as 

 did a rather less specific esterase inhibitor like rhodamine. 



The situation in red blood cells may be similar. Greig, Faulkner, and 

 Mayberry (1953) have shown that active transport or uptake of K+ can be 

 promoted by suitable metabolic energy sources and also by supply of acetyl- 

 choline in their absence, but that this transport is inhibited by physostigmine. 

 There is some doubt regarding details of this situation as the concentration 

 of physostigmine for inhibition of salt uptake is rather high, 2xlO'^M, 

 while the isolated enzyme system is inhibited at 10^^ M (Strickland and 

 Thompson, 1955). It is, however, of interest that concentrations of this 

 order, 10"^ M lAA, cause roughly 50-60 per cent inhibition of uptake of 

 NaCl from 0-02 M solutions by discs of beetroot tissue. Similar concentrations 

 of lAA cause pronounced exosmosis of salts from coleoptile segments as 

 shown by increase in conductivity of the water in which they are floating and 

 also inhibition of salt uptake. 



Rhodamine at a concentration of 10 mg/1. causes an 80 per cent reduction 

 of NaCl uptake by beetroot discs. 



(v) These facts do not, of course, show that salt uptake by plant tissue is 

 mediated by an acetylcholine-esterase system. It is, moreover, necessary to 

 show first the presence of an active choline esterase and here it is somewhat 

 difficult to obtain really precise evidence. Discs of beetroot and roots of oat 

 and of bean do cause marked decrease in pH of acetylcholine chloride 

 supplied at 2xlO~^M at pH 7-3, but this could be explained as due to 

 exosmosis of ions from the tissue. Parallel to this there is loss of acetylcholine 

 from the external solution as determined colorimetrically after conversion 

 to acethydroxamic acid ; this loss, however, could be due to absorption of the 

 acetylcholine chloride by the tissue. It is clearly necessary to obtain the 

 isolated enzyme system before one can have confidence in its presence or 

 consider very seriously its role in effecting ion uptake. 



Dr. Bell of the Zoology Department of King's College has kindly carried 

 out the histochemical test using acetylthiocholine chloride in a copper- 

 glycine buffer. Crystals were formed at the surfaces of root hairs of broad 

 bean, Viciafaba, but in material simultaneously treated with physostigmine or 



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