Louis B. Flexner

Memory in Mice as Affected by Intracerebral Puromycin

The antibiotic, puromycin, caused loss of memory of avoidance discrimination learning in mice when injected intracerebrally. Bilateral injections of puromycin involving the hippocampi and adjacent temporal cortices caused loss of short-term memory; consistent loss of longer-term memory required injections involving, in addition, most of the remaining cortices. Spread of the effective memory trace from the temporal-hippocampal areas to wide areas of the cortices appears to require 3 to 6 days, depending upon the individual animal. Recent reversal learning was lost while longer-term initial learning was retained after bilateral injections into the hippocampal-temporal areas.

INHIBITION OF PROTEIN SYNTHESIS IN BRAIN AND LEARNING AND MEMORY FOLLOWING PUROMYCIN

ON the assumption that learning and memory may have a biochemical basis, the suggestion has become increasingly frequent during recent years that they may depend in some way on macromolecules such as nucleic acid or protein. The discovery by YARMOLINSKY and DE LA HABA (1960) that puromycin produces profound inhibition of protein synthesis in a cell-free system and the later demonstration that it efficiently suppresses protein synthesis in vivo (GORSKI, AIZAWA and MUELLER, 1961) led us to investigate its effect on the central nervous system. This report deals primarily with the substantial suppression of protein synthesis which has been produced in the brains of mice with puromycin, and with the performance of these mice in tests of their ability to learn and to retain memory of the learning experience.

The comparative physiology of placental transfer

Abstract The transfer of sodium across the placentas of six different animals at progressive stages of the gestation period has been studied. The rate of transfer of sodium per gram of placenta increases in each animal as gestation proceeds until just before term, at which time there is a sharp decrease. The variation in rate of transfer across the placentas has been correlated with morphologic changes in the placenta. The animals have been chosen to give at least one representative of each of Grossers morphologic types of placentas. The rates of transfer of sodium across unit weights of these four types of placentas have been found to depend upon the morphologic structure of the placenta: the smaller the number of tissue layers placed between maternal and fetal circulations, the greater the rate of transfer across a unit weight of the placenta. A correlation was found to exist between the supply of sodium transferred to a unit weight of fetus and the rate at which that unit weight of fetus was growing. This constant observation in all of the animals studied leads us to propose the hypothesis that the underlying principle of placental function is that the rate at which a physiologic substance is transferred to the fetus shall parallel the relative growth rate of the fetus.

LOSS OF MEMORY AS RELATED TO INHIBITION OF CEREBRAL PROTEIN SYNTHESIS

IT was previously reported (FLEXNER, FLEXNER and STELLAR, 1963) that the effective locus of the memory trace of simple maze learning in mice appears to spread from the hippocampi and temporal cortices to remaining areas of the neocortex in from 3-6 days after the learning experience. Memory before this period of transition has been called short-term or recent memory; after this period, longer-term memory. Recent memory can be consistently destroyed by bilateral temporal injections of puromycin which have been found to inhibit protein synthesis in the hippocampus and temporal cortex by at least 80 per cent for 8-10 hr. (FLEXNER, FLEXNER, OBERTS and DE LA HABA, 1964). Bilateral temporal injections are, however, without effect on longer-term memory, the loss of which depends upon the use of combined bilateral temporal plus ventricular plus frontal injections (FLEXNER et al., 1963). The present report is concerned with the inhibitory effects of these combined bilateral injections on cerebral protein synthesis. In addition, we have measured the rate of cerebral protein synthesis in the presence of several substances which are related to puromycin but which have no effect on memory.

Memory and cerebral protein synthesis in mice as affected by graded amounts of puromycin

Abstract The effects of intracerebral injections of graded amounts of puromycin on memory and on degree and duration of inhibition of cerebral protein synthesis have been studied in the mouse. As the concentration of the antibiotic was decreased it became progressively less effective in its behavioral and biochemical effects so that memory was retained in an increasing proportion of animals as the effect on protein synthesis diminished.

The rate of renewal in woman of the water and sodium of the amniotic fluid as determined by tracer techniques

Abstract The rate of passage of water and sodium from the maternal circulation to the amniotic fluid has been measured with heavy water and radioactive sodium as the tracer substances. The water of the fluid is completely replaced on the average once every 2.9 hours; this considerable rate of turnover is at variance with the concept that the amniotic fluid is stagnant. The rate of transfer of water is about five times more rapid than that of sodium.

BIOCHEMICAL AND PHYSIOLOGICAL DIFFERENTIATION DURING MORPHOGENESIS‐XXIII. FURTHER OBSERVATIONS RELATING TO THE SYNTHESIS OF AMINO ACIDS AND PROTEINS BY THE CEREBRAL CORTEX AND LIVER OF THE MOUSE*

THE observations presented here in part supplement previous ones concerning the newborn mouse (FLEXNER, FLEXNER and ROBERTS, 1958). In major part, however, they relate to the adult animal. Both cerebral cortex and liver have been studied to obtain estimates of the following quantities: (a) the rate at which blood supplies glutamic acid to these tissues; (b) the rate at which the carbon of glucose is incorporated into several non-essential amino acids of the tissues; (c) the rate at which certain essential amino acids are supplied to the tissues by the blood; and (d) the rate at which these amino acids in the tissues are incorporated into protein. It has been our interest to study the effect of growth and maturation on these quantities. The findings of GAITONDE and RICHTER (1956) with methionine and of LAJTHA, FURST, GERSTEIN and WAELSCH (1957) and LAJTHA, FURST and WAELSCH (1957) with lysine leave no doubt that amino acids are incorporated at a substantial rate into the proteins of the adult as well as the newborn brain.

Lactic dehydrogenases of the developing cerebral cortex and liver of the mouse and guinea pig

Abstract Using cellulose ion exchangers and starch gel electrophoresis, four components of LDH have been found in the cerebral cortex of mouse and guinea pig. The several components tested could also be distinguished by their ratios of rates of reduction of DPN and two of its analogs. These components apparently each contribute to a different degree to the increase in LDH activity of the cortex which occurs during development. Maturation of the liver is accompanied by a reduction in the number of components of LDH in both mouse and guinea pig. With the liver of the mouse, peaks of LDH activity in the chromatograms which corresponded in their position with those from cerebral cortex also gave the same ratios of reduction of DPN and its analogs.

Dose-response relationships in attenuation of puromycin-induced amnesia by neurohypophyseal peptides

Intracerebral injections of puromycin one day after training of mice in a Y-maze cause amnesia when the animals are tested 7 days later. This amnesia was shown to be attenuated by various neurohypophyseal hormones, analogs and fragments, administered subcutaneously immediately after training. Dose-response relationships have been obtained for the attenuation of puromycin-induced amnesia in mice by selected neurohypophyseal peptides. All of the compounds tested reduce the amnesia in a dose-related way, suggesting that these peptides may interact with specific receptors to induce their central effect. Among the peptides studied the two most potent--i.e., those that cause substantial retention of memory at the lowest doses--are the neurohypophyseal hormone arginine vasopressin and Z-prolyl-leucyl-glycinamide (Z-MIF).

Intracerebral Saline: Effect on Memory of Trained Mice Treated with Puromycin

It was shown that puromycin administered to mice 1 or more days after maze-learning blocks expression of memory; the blockage can be removed by intracerebral injections of saline. We present evidence that intracerebral injections of saline are relatively ineffective in restoring memory when puromycin is administered either before or immediately after training; in these two situations puromycin appears to interfere with consolidation of memory.