Abstract. Before addition of cAMP, Dictyostelum amoebae rapidly translocating in buffer are elongate, exhibit expansion zones primarily at the anterior end and filamentous actin (F-actin) localization primarily in the anterior pseudopodia. Intracellular particle movement is primarily in the anterior direction, and the average rate of particle movement is roughly five times the rate of cellular translocation. Within seconds after the addition of 10 -6 M cAMP, there is a dramatic suppression of cellular translocation, an inhibition of pseudopod formation, a freeze in cellular morphology, a dramatic depression in intracellular particle movement, loss of F-actin localization in pseudopodia concomitant with relocalization of F-actin in the general cytoplasmic cortex under the plasma membrane, and a doubling of F-actin content. After 10 s, expansion zones are again visible at the cell perimeter, but they no longer are localized in the original anterior portion of the cell. There is a slight rebound in particle movement after 10 s, but particles with persistent tracks now show no directionality towards the original anterior portion of the cell, as they did before cAMP addition. Finally, in parallel with the resumption of peripheral expansion and the small rebound in particle movement, there is a decrease in total cellular F-actin to the untreated level. The pattern of microtubule organization is unaffected by the addition of cAMP.URING aggregation in the cellular slime mold Dictyostelium discoideum, cells in the center of an aggregation territory release the chemoattractant cAMP in a pulsatile fashion (3,30,40). Cells peripheral to the center respond by relaying the signal outwardly (6, 10) and by moving in a directed fashion towards the aggregation center (1). Because of the pulsatile nature of the original signal and the relay system, cAMP moves outwardly through the territory as a nondissipating wave. The extracellular cAMP signal is mediated by a cell surface receptor that is a member of the beta-adrenergic receptor family (15) and interacts with G proteins (16, 31).To investigate the sequence of receptor-mediated biochemical events in the cAMP response, the standard protocol has been the rapid addition of cAMP to chemotactically responsive cells suspended in or perfused with buffer. Although the increase in cAMP in the cellular environment under these experimental conditions occurs at a far faster rate than during the front of a natural wave (35, 44), a number of rapid physiological responses have been demonstrated which probably play integral roles in cAMP-mediated chemotaxis. These changes include (a) the synthesis and release of cAMP (5) Recently, the behavioral responses of cells to a rapid increase in cAMP were assessed. It was demonstrated that the rapid addition of cAMP to the peak concentration of the natural wave (10-6 M) results in an immediate decrease in velocity measured by centroid translocation (35,43,44,46), an increase in directional change (44), an increase in roundness (13,35,44), and a decrease...

When developing amebae of Dictyostelium discoideum are treated with constant concentrations of cAMP above 10(-8)M, the average rate of motility is depressed, with maximum inhibition at roughly 10(-6)M. It is demonstrated that shifting the concentration of cAMP from 0 M to concentrations ranging from 10(-8) to 10(-6)M in a perfusion chamber results in the immediate inhibition of motility. After shifting from 0 M to 10(-8) or 10(-7)M, the rate of cell motility remains low, then rebounds to a higher level, exhibiting a standard adaptation response. No adaptation is exhibited after a shift from 0 M to 10(-6)M, a concentration resulting in maximum inhibition. It is demonstrated that the level of inhibition and the extent of the adaptation period are dependent upon the concentration of cAMP after the shift, and that submaximal inhibition is additive. The characteristics of adaptation in this motility response are very similar to the characteristics of adaptation for the relay system and phosphorylation of the putative cAMP receptor.