Changes of cyanide content and linamarase activity in wounded cassava roots

When cassava (Manihot esculenta Crantz) root was cut into blocks and incubated under laboratory conditions, the blocks showed more widespread and more even symptoms of physiological deterioration than those under natural conditions. Thus, the tissue block system has potential for biochemical studies of natural deterioration of cassava root. The changes in cyanide content and linamarase (linamarin β-d-glucoside glucohydrolase; EC 3.2.1.21) activity in various tissues during physiological deterioration were investigated. Total cyanide content increased in all parts of block tissue after 3-day incubation. The degree of increase in cyanide was most pronounced in white parenchymal tissue, 2 to 3 millimeters thick, next to the cortex (A-part tissue), where no physiological symptoms appeared. On the other hand, linamarase activity was decreased in all parts of block tissue after a 3-day incubation. A time course analysis of A-part tissue indicated a clear reciprocal relationship between changes in total cyanide and linamarase activity; total cyanide increased, while linamarase activity decreased. Free cyanide constituted a very small portion of the total cyanide and did not change markedly.     

Ionic channels and nerve membrane constituents. Tetrodotoxin-like interaction of saxitoxin with cholesterol monolayers

Saxitoxin (STX) and tetrodotoxin (TTX) have the same striking property of blocking the Na⁺ channels in the axolemma. Experiments with nerve plasma membrane components of the squid Dosidicus gigas have shown that TTX interacts with cholesterol monolayers. Similar experiments were carried out with STX. The effect of STX on the surface pressure-area diagrams of lipid monolayers and on the fluorescence emission spectra of sonicated nerve membranes was studied. The results indicate a TTX-like interaction of STX with cholesterol monolayers. The expansion of the monolayers caused by 10^-6 M STX was 2.2 A²/cholesterol molecule at 25°C. From surface pressure measurements at constant cholesterol area (39 A²/molecule) in media with various STX concentrations, it was calculated that the STX/cholesterol surface concentration ratio is 0.54. The apparent dissociation constant of the STX-cholesterol monolayer complex is 4.0 x 10^-7 M. The STX/cholesterol ratio and the apparent dissociation constant are similar to those determined for TTX. The presence of other lipids in the monolayers affects the STX-cholesterol association. The interactions of STX and TTX with cholesterol monolayers suggest (a) that cholesterol molecules may be part of the nerve membrane Na⁺ channels, or (b) that the toxin receptor at the nerve membrane shares similar chemical features with the cholesterol monolayers.     

ADENOSINE TRIPHOSPHATASE ACTIVITY IN THE MEMBRANES OF THE SQUID NERVE FIBER

This investigation deals with the localization of sites of ATPase activity, especially of transport ATPase, in nerve fibers of the squid Doryteuthis plei, at the subcellular level. Splitting of ATP liberates inorganic phosphate which reacts with lead to form a precipitate in the tissue. The reaction was made on nerve fibers fixed with glutaraldehyde. Frozen slices were incubated in Wachstein-Meisel medium containing ATP and Pb(NO₃)₂. Deposits of reaction product were found in the axolemma (towards its axoplasmic side), Schwann cell membranes (mainly at the channels crossing the layer), and mitochondria. Control experiments revealed that no deposits were observed in nerve fibers fixed in osmium tetroxide prior to incubation in the medium containing ATP, or in nerve fibers incubated without substrate or with adenosine monophosphate, adenosine diphosphate, glycerophosphate, or guanosine triphosphate as substrate. For evaluation of transport ATPase activity, these findings were compared with results obtained with nerve fibers treated with G-strophanthin or K-strophanthoside before or after glutaraldehyde fixation. The cardiac glycosides produced a disappearance or diminution of the deposits. The largest inhibitory effect was observed in the axolemma. The findings indicate that the highest ATPase activity is localized in the axolemma and may be due primarily to transport ATPase.     

Water diffusion under osmotic gradients in frog gastric mucosa

The asymmetry of the osmotic response of the frog gastric mucosa has been further analyzed by studying the effect of external tonicity changes on the water diffusion fluxes. Hyperosmotic solution at the serosal surface does not affect the water diffusion fluxes. Hyperosmotic solution at the mucosal surface, with isosmotic solution at the serosal surface, significantly reduces (P<0.001) the serosal-to-mucosal and the mucosal-to-serosal water diffusion. An increment in the restriction offered by the mucosa to water diffusion by effect of hypertonicity at the mucosal surface is proposed.     

Non-electrolyte diffusion across the frog gastric mucosa under osmotic gradients

Serosal-to-mucosal and mucosal-to-serosal diffusion of ¹⁴C-labelled inulin, sucrose, erythritol and propionamide was compared with ³HHO diffusion in mucosae incubated with isosmotic solutions at both surfaces, as well as isosmotic solution at serosal surface and hyperosmotic solution at the mucosal surface. The use of a hyperosmotic solution at the mucosal surface significantly increases unidirectional diffusion fluxes of inulin and of sucrose. To a nonsignificant extent, it affects the fluxes of erythritol and propionamide and significantly reduces the ³HHO diffusion. A size increment of the diffusion path utilized by the larger molecules is proposed.