195

the rats received the drug. More import~t~y, Krank and Bennett showed that this response is transferred to morphine, even when morphine is given at a dose that normally should induce a decrease, rather than an increase, in activity5.

The impBcations of these find- ings for human addictive behavior are obvious, particularly in relation to environment or contextual effects switches.

and drug

TIPS - June 1988 fVol. 91

How environment affects conditioned drug tolerance Repeated drug administration in a given stimulus context may induce the development of a con- ditioned response, and this response may occur in anticipa- tion of I, and interact with *, the regular ‘unconditioned’ effect of the drug. Very often, the con- ditioned response is a ‘compensa- tory’ response; i.e. one that con- sists of an effect opposite to the one that would be expected from the drug (e.g. an increase instead of a decrease of locomotor activ- ity).

Siegel and his co-workers have expanded these early observations into a conditioned-reflex model of drug tolerance, and have indeed demonstrated a role of condition- ing in the development of toler- ance for the analgesic effect of morphine, and for the hypother- mic effects of ethanol, among other drugs’. The problem is of practical as well as theoretical importance. A lot of the drug- seeking behavior of drug abusers

is said to be conditioned, partly to the en~~nment and/or the social context; and it is common for drug abusers to switch from a given drug or drug combination to another (usually cheaper or more socially acceptable) in order to procure the same ‘high’. Many die in this quest, either because they succeed or because they don’t.

Two papers bearing on this point have recently appeared. Pal- letta and Wagner* reported on the development of context-specific tolerance to both the analgesic and the activity-depressant effects of morphine in rats: conditioned ‘cornpen~to~~ responses occur- red only in the activity measure. Krank and Bennett5 reported on the development of context- specific conditioning to the locomotor-stimulant effect of (+)- amphetamine, also in rats: pre- and post-drug activity levels increased over a number of daily sessions but only in the box where

IVAN IZQUIERDO

Centro de Men&a, Deparfamento de Biu- quimica, lnsti~~~ de B~octe~cius, UFRGS, 90049 Porto Alegre, RS, Brazil.

References 1 Pavlov, 1. P. (1927) Conditioned Reflexes,

Oxford University Press 2 Bykov, K. M. (1959) The Cerebral Cortex

and the Inter& Organs, USSR Foreign Languages Publishing House

3 Siegel, S. and MacRae, J. (1984) Trends Neurosci. 7,140-143

4 PaIk?tta, M. S. and Wagner, A. R. (1986) Behuv. Neurosei. 1UO. 611623

5 Krank, M. D. and. Bennett, D. (1987) Behav. Neural Biol. 48,422-433

Controlled expiation of the cell interior by pore-forming proteins Ex~riment~ analysis of intracel- lular processes is hampered by the limiting plasma membrane. Thus data from intact cells often pro- vide only indirect evidence on the nature and regulation of intracel- lular events. Isolated enzymes and purified subcellular organelles can be characterized biochemically but this gives no information about their cellular function.

In contrast, permeabilization of the plasma membrane by appro- priate techniques allows access to, and control over, the cytosol while leaving the intracellular organiza- tion intact. Such permeabilized cell preparations permit study of the regulation of exocytosis in secretory cells, a process which is contralied by intracellular events and involves the fusion of secre- tory vesicles with the plasma membrane. The chain of events leading to exocytosis and its mod-

ulation are still not completely understood.

Techniques for membrane pe~eabil~ation

Exocytosis can be studied in permeabilized secretory cells, pro- vided that neither the vesicle membrane nor the plasma mem- brane has been disturbed by the pe~eab~i~tion procedure. Per- meabilization of secretory cells has hitherto been performed by physical or chemical techniques.

Iiigh voltage discharge, first used with adrenal medullary chro- maffin cells’ allows dialysis of small molecules from the cell interior, whereas soluble cytoplas- mic proteins remain within the cell*. Electrical permeabilization is especially suitable for cells in sus- pension.

The whole cell patch clamp technique, which records the

increase in capacitance of the plasma membrane during exocy- tosis, is a method for permeabili- zation of single cells. It allows the introduction of not only small molecules but also high molecular weight proteins into the cell. Furthe~ore it is possible to analyse the fusion of single vesi- cles with the plasma membrane3.

Detergents can be used to per- meabilize cultured cells in sus- pension or attached to culture plates. In the past, saponin and digitonin have been used for the permeabilization of adrenal medullary chromaffin and other secretory cellsz+j Iiowever, detergents may destroy intracellu- lar membranes2f7.

To overcome these and other difficulties inherent in the pre- vious techniques we have employed bacterial toxins for the stable, selective and prolonged permeabilization of the plasma membrane of secretory cells’-“. Two pore-forming proteins, ix- toxin from Staphylococcus aureus and streptolysin 0 from p- hemolytic streptococci, permit the pe~eabilization of secretory cells