Cyclic nucleotides in glutamate chemosensory signal transduction of Paramecium

Journal of Cell Science, Oct 1997

W.Q. Yang, C. Braun, H. Plattner, J. Purvee, J.L. Van Houten

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Cyclic nucleotides in glutamate chemosensory signal transduction of Paramecium

W. Q. Yang 1 2 C. Braun 0 1 H. Plattner 0 1 J. Purvee 1 2 J. L. Van Houten 1 2 0 Department of Biology, Universitat Konstanz , Konstanz , Germany 1 Key words: Paramecium , Cyclic AMP, Cyclic GMP, Glutamate, Chemoreceptor, IMP 2 Department of Biology, University of Vermont , Burlington, VT *Author for correspondence - Glutamate is an attractant stimulus to Paramecium tetraurelia. It causes a hyperpolarization of the cell and smooth, relatively fast swimming that is characteristic of hyperpolarizing stimuli. We show here that by 1-30 seconds of stimulation, glutamate increases intracellular cAMP. Interestingly, other attractant stimuli, such as acetate and NH4Cl, that similarly hyperpolarize the cell do not induce an increase in cyclic AMP observable at 30 seconds. In order to determine whether the changes in cyclic AMP could be rapid enough to participate in stimulation as compared to slower processes such as adaptation, rapid kinetic measurements of cyclic AMP were made on whole cells by quenched-flow. We found that, in cells stimulated with glutamate, intracellular cyclic AMP increases by 30 mseconds and peaks at about sevenfold over basal levels by Glutamate has been recognized as an important intercellular signal molecule for neurotransmission (Tanabe et al., 1992; Wo and Oswald, 1995; Nakanishi, 1992). Perhaps not as well appreciated is that glutamate also serves as an important environmental cue for many different organisms. Lobsters (Carr and Derby, 1986; Fine-Levy et al., 1987), fish (Caprio et al., 1993), and mammals including humans (Yamaguchi, 1987) can detect, that is, taste and/or smell, glutamate among other amino acids. Mammalian taste has the added interesting aspect of synergism of glutamate with 5 ribonucleotides, known as umami taste (Ugawa and Kurihara, 1994; Faurion, 1991). Therefore, it is not unusual that paramecia detect and are attracted to glutamate, which could very well indicate that bacteria, their food, are at hand. Preston and Usherwood (1988) described attraction to glutamate and specific binding sites on cilia. Likewise, in our studies, glutamate is an attractant to P. tetraurelia, albeit at higher concentrations than used by Preston. The attractant responses are consistent and robust, as shown in this paper. Attractants of Paramecium tetraurelia tend to be small molecules that probably indicate the presence of bacteria. Not only glutamate, but also acetate, extracellular cyclic AMP, biotin, folate, and NH4Cl are among the stimuli that we have 200 mseconds. Cyclic GMP does not change relative to basal levels over rapid or slower time courses of glutamate stimulation. An antagonist of glutamate, IMP, depolarizes the cells and decreases intracellular cyclic AMP by approx. 50% and slightly increases cyclic GMP. Results of behavioral tests of cells treated with protein kinase inhibitors also suggest that cyclic AMP is part of the signal transduction pathway for glutamate, but not for other attractant stimuli. These studies are the first demonstration of a possible role for cyclic nucleotide second messengers in an attractant chemosensory transduction pathway in Paramecium. identified (Van Houten, 1978; Van Houten and Preston, 1987; Bell and J. L. Van Houten, unpublished observations). They have in common that they all hyperpolarize cells and cause relatively fast and smooth swimming. Such swimming behavior, with adaptation, causes the accumulation of cells by a biased random walk (Van Houten, 1990). There are at least three different signal transduction pathways, or at least alternate versions of pathways, and the attractant stimuli acetate, NH4Cl, and glutamate each stimulate a different pathway. The pathways epitomized by acetate and glutamate are cell surface receptor-mediated, and that epitomized by NH4Cl involves intracellular pH modulation probably without receptor involvement. Hyperpolarization by altering membrane potential (Vm) through lowering extracellular K levels has long been demonstrated to elevate Paramecium cyclic AMP levels, and both hyperpolarization and cyclic AMP correlate with increased ciliary beating and rapid swimming (Machemer, 1989; Pech, 1995). Here we show that hyperpolarization of the cells induced not only by altering extracellular K, but also by receptor-mediated processes raises intracellular cAMP levels. What is even more interesting is that this second messenger response is specific to the glutamate pathway; other attractant stimuli that hyperpolarize do not appear to elevate cAMP. We also report here that protein kinase inhibitors interfere specifically with attraction to glutamate, which is consistent with the demonstration in this paper that glutamate alone among the attractants tested increases intracellular cyclic AMP. Additionally, we demonstrate that the increase in cyclic AMP occurs rapidly enough to be consistent with a role in the signal transduction pathway as opposed to or perhaps in addition to a role in slower processes such (...truncated)


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W.Q. Yang, C. Braun, H. Plattner, J. Purvee, J.L. Van Houten. Cyclic nucleotides in glutamate chemosensory signal transduction of Paramecium, Journal of Cell Science, 1997, pp. 2567-2572, 110/20,