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)