Malonate and Cyanide Insensitivity in Relation to Respiratory Compensation in Potato Slices.

The respiration of a variety of plant tissues has been shown to be ostensibly unaffected by malonate or by cyanide. Inhibitor insensitivity may be a characteristic of the tissue in its natural condition, such as the cyanide insensitivity of the basal respiration of wheat roots (13) or of the respiration of A4rwn spadices (6, cf. 2). On the other hand, insensitivity may develop when respiratory behavior is altered experimentally, as is the case with cyanide insensitivity of slices of a variety of plant storage organs which have been aged (4, 14, 23). Parenthetically, normally inhibitor-resistant tissue may display considerable sensitivity elicited by experimental treatment or simply by time. Thus the malonate-resistant, cyanide-sensitive respiration of fresh potato slices gives way with time to a predominantlv malonate-sensitive, cyanide-resistant respiration (8. 21), while a cyanide-sensitive salt respiration may be evoked in barley roots or carrot slices, for example, superimposed upon a basal cyanide-resistant metabolism ( 16). In any event, insensitivity to a respiratory inhibitor raises the fundamental question of whether resistance reflects the operation of a respiratory path unsusceptible to inhibitor action, or whether the inhibitor in fact exerts its accustomed effect, with the result that the inhibition educes alternative compensating respiratory activity. There are several indications that respiratory compensation occurs. On the one hand, respirationdependent physiological processes remain sensitive to terminal oxidase inhibitors while the gross respiration becomes resistant (3). On the other hand, direct evidence suggests that electron transport proceeds through an alternative chain of electron carriers when the ostensibly normal electron path is blocked by cyanide, among other inhibitors (1, 4, 5, 24). In the latter connection, diversion of electron transport is frequently marked by respiratory stimulation (11). Unfortunately a demonstrated capacity on the part of a given tissue or organelle to use alternative respiration pathways gives no answer to the basic question of which path, in the absence of inhibitor, is operative in vivo. Where the alternatives apply to electron paths a determination may be sought spectrophotometrically. Although a likely answer has been obtained in the case of the Aroidi spadix (25), an unambiguous assessment of electron pathway in intact tissue by spectrophotometric means is a difficult task (12, 13). Where it is likely that the carbon path of respiration differs in a given tissue under different conditions, for example in malonatesensitive as compared with malonate-resistant potato slices, there is the need for a means to evaluate the nature and prevalence of alternative carbon paths in the presence and absence of inhibitor, if the question raised at the outset is to be answered. Even where electron pathways are primarily at issue there remains a question regarding the extent to which curtailment of a given electron path may affect associated carbon-path metabolism. Experimental Rationale. Considerable cyanide resistance develops in potato slices in the course of a day's incubation (14, 23). Although the respiration rise in potato slices with aging is normally largely malonate-sensitive., the developed respiratory increment is virtually malonate-resistant when disks are aged in bicarbonate solution in an atmosphere of 10 % CO, in aiir (8, 9, cf. 7). Thu's by aging potato slices either normally or in a bicarbonate-CO. environment, slices of high respiratory activity are obtained, resistant to cyanide and malonate respectively. In the experiments which follow, the effect of malonate is determined on the course and extent of C14 incorporation into a variety of endogenous metabolites following the presentation of glucose-U-C14 in tracer quantities. Special attention is paid to components of the tricarboxylic acid cycle, it having been shown that the tricarboxylic acid cycle constitutes much of the respiratory activity of normallv aged slices (8, 10). The rationale is simply that malonate should have no effect on label incorporation and distribution in malonate-insensitive tissue if the metabolic path is truly indifferent to malonate, but should have an effect resembling that in sensitive tissue if malonate represses the normally operative path and thereby evokes an alternative path. The effect of cyanide is examined in the same way.