J. L. Van Houten

Computer simulation of Paramecium chemokinesis behavior

Abstract A computer program was designed to simulate the distribution of paramecia in a T-maze assay for chemo-accumulation and dispersal. Simulated values of chemokinesis are compared to experimental values for normal and mutant paramecia. The roles of components of swimming behavior (turning frequency and swimming speed), adaptation, and reaction at the border of solutions are examined.

Chemoreception in Paramecium tetraurelia: acetate and folate-induced membrane hyperpolarization.

SummaryAcetic and folic acids hyperpolarize the membrane potential ofParamecium tetraurelia in a concentration-dependent manner. The membrane responses are accompanied by small changes in cell resistance, and are significantly reduced by increasing extracellular cation concentrations, suggesting that the attractants bring about the membrane potential change by increasing cell permeability to cations. The inability to show a reversal potential for the hyperpolarization to attractants suggests that the effects of cations on the response are non-specific, however. The possible roles of Ca++, K+, and Na+ in the attractant-induced responses were further investigated by applying acetate and folate to cells with genetic defects in specific ion conductances, by collapsing the driving forces for these ions, and by testing the effects of ion channel blockers on the responses. These studies suggest that the membrane responses to attractants are not due to the direct effects of increased or decreased membrane permeability to cations.Attempts to block the acetate and folate-induced hyperpolarization by collapsing surface potential or using a mutant with reduced surface charge were inconclusive, as were studies on the possible role of attractant transport in the membrane responses.We hypothesize that the membrane hyperpolarization may be due to either the indirect effects of increased calcium permeability, to extrusion of calcium through activation of a calcium pump, or to a proton efflux.

Ca2+ transport and chemoreception in Paramecium

Intracellular Ca2+ levels in Paramecium must be tightly controlled, yet little is understood about the mechanisms of control. We describe here indirect evidence that a phosphoenzyme intermediate is the calmodulin-regulated plasma membrane Ca2+ pump and that a Ca2+-ATPase activity in pellicles (the complex of cell body surface membranes) is the enzyme correlate of the plasma membrane pump protein. A change in Ca2+ pump activity has been implicated in the chemoresponse of paramecia to some attractant stimuli. Indirect support for this is demonstrated using mutants with different modifications of calmodulin to correlate defects in chemoresponse with altered Ca2+ homeostasis and pump activity.

Relationship of folate binding to chemoreception inParamecium

SummaryParamecia respond to some soluble chemicals in their environment. Cells change their swimming pattern to accumulate in or disperse from these chemicals. Attractants of one class, which includes the folate anion, hyperpolarize the cell relative to the membrane potential in control solution (Van Houten 1979). The mechanism of transduction of the chemical cue (folate) into its characteristic electrical cue is unknown. This study investigates the role of specific binding in this transduction. While the role of uptake cannot be unequivocally established (DiNallo et al. 1980), the role of binding in chemoreception is more clear and appears to be a necessary step in the chemosensory transduction pathway. Evidence for this comes from concomitant loss of saturable folate binding with loss of chemoresponse both in mutant d4-534 and assays of binding and chemoresponse in the presence of cyclic AMP. Normal folate binding toParamecium tetraurelia is characterized by a Kd of approximately 29 μM and a binding site number of approximately 6.7 × 10−11 mol/mg protein.

Intracellular pH and chemoresponse to NH4+ in Paramecium

Paramecium are attracted to ammonium chloride solutions relative to sodium chloride control solutions, but little is known about the mechanisms by which attraction is evoked. A known effect of ammonium solutions in other cell types is an alteration of intracellular pH. We show here that intracellular pH is elevated upon initial exposure to 5 mM NH4Cl, but appears to decline within 10 minutes, both in wild type cells and in two mutants which do not show sustained attraction to NH4Cl using the standard behavioral assay, the T-maze. We also present quantitative values of swimming parameters that underlie the response to NH4Cl.

Localization of the chemoreceptive properties of the surface membrane of paramecium tetraurelia

SummaryThe plasma membrane ofParamecium tetraurelia comprises two morphologically distinct components; a membrane that encloses the cell body and a ciliary membrane. In order to investigate the relative contributions of the two membranes to attractant-induced membrane potential changes, cells were deciliated with ethanol and their subsequent responses to attractants examined.Deciliation did not significantly affect the magnitude of the hyperpolarizations evoked by acetic or lactic acids, and had no effect on the concentration dependence of responses to folic acid. We conclude that the components necessary for detection and response to attractants are not exclusive to the ciliary membrane ofP. tetraurelia. Deciliation ofParamecium concomitantly permits localized chemical stimuli to be applied directly to the cell surface in the absence of strong fluid currents that are generated by the activity of the locomotory organelles. By systematically applying K2 folate to a number of sites on the cell surface, it has been possible to demonstrate an anterior-posterior gradient of chemosensitivity on the cell body ofP. tetraurelia.

BIOTIN CHEMORESPONSE IN PARAMECIUM

Paramecium tetraurelia locate their␣foodsource by detecting bacterial metabolites and altering swimming behavior to congregate near bacterial populations on which they feed. Several attractants, such as folate, glutamate, cAMP and acetate have been identified and various aspects of chemoreception, signal transduction and effector mechanisms have been described. Here we characterize the Paramecium chemoresponse to biotin. An essential enzymatic cofactor in all cells, biotin is secreted by a large number of bacterial species during growth phase. P. tetraurelia are strongly attracted to biotin with a half-maximal behavioral response at 0.3 mmol · 1−1 in T-maze assays. Physiological recordings from whole cells show that cells hyperpolarize in a concentration-dependent manner in biotin. Whole-cell binding assays utilizing 3H-biotin identify a saturable and specific binding site with an apparent dissociation constant of 0.4 mmol · l−1. The biotin analogs desthiobiotin and biotin methyl ester are also strong attractants. Diaminobiotin fails to attract P. tetraurelia at 1 mmol · l−1, but does interfere with the biotin chemoresponse and displaces 3H-biotin from whole cells. We hypothesize that the keto group and/or fidelity of the ureido ring of biotin are necessary for biotin chemoresponse.

Meckelin guides orientation of basal bodies along the striated rootlet

Objective Meckelin (MKS3) functions in ciliogenesis and ciliary gating. MKS3 appears to have similar functions in Paramecium tetraurelia, i.e. FLAG-MKS3 is found associated slightly above each basal body and RNAi for MKS3 leads to loss of cilia. RNAi for MKS3 also leads to the disorganization of rows of basal bodies that run from anterior to posterior. In areas of misalignments, basal bodies with their post ciliary and transverse rootlets are found out of their expected rows. However, the rootlets are attached to the basal bodies at the expected angles relative to each other. We propose that MKS3 guides new basal bodies as they move toward the anterior of the cell along the striated rootlet (SR) of the parent basal body. Basal bodies without MKS3 lose their interactions with the parent’s SR. Without this guide to maintain orientation, new basal bodies migrate off the expected line and, when they form their SRs, these too cannot project toward the anterior as expected.

Chemical Senses: Protozoa

Protozoans are solitary cells that nonetheless can be social; they use chemical cues to congregate together in areas of food or mates or to become the focal point for differentiation into cysts, to weather adverse environmental conditions. Protozoans encompass many forms, including cells covered with cilia, cells that propel themselves with one or two flagella, and cells that crawl. What they all have in common is the ability to sense chemical stimuli in their environment and move in response to these stimuli. We focus here on protozoan chemical senses and further focus on a few examples of ciliates and social amoebae.