The overall goal of my research program is to better understand the pharmacology, biophysics, structure and function of neurotransmitter receptors that respond to the amino acid glutamate. These receptors exist in the nervous systems of both vertebrate and invertebrate animals. The mammalian receptors are the best understood, and are classified into two general categories: metabotropic (receptor activation linked to activation of G proteins) and ionotropic (receptor activation directly coupled to ion channels). My current research focuses primarily on the characterization of glutamate receptors of involved in the feeding behavior of the pond snail,Helisoma trivolvis
Glutamate receptors involved in feeding in pond snails
The pond snail, Helisoma trivolvis, has a relatively simple central nervous system consisting of several ganglia (groupings of neurons) linked together in two rings. In this system neuronal activity can be correlated closely with the behaviors of the animal. Glutamate is used as a neurotransmitter influencing the feeding behavior of these snails. Both glutamate-containing and glutamate-responsive neurons have been identified in the buccal ganglia that control feeding (Quinlan et al., 1995). Of particular interest is the finding that glutamate both excites and inhibits motor neurons regulated by the central pattern generators that control feeding behavior. It is not yet known what types of glutamate receptors are present in this system. The excitatory receptors are probably similar to a type cloned from a closely related pond snail, Lymnaea stagnalis, and have 44-48 % sequence homology with the mammalian kainate receptor subunits. The activity of the receptor from Lymnaea is blocked by the mammalian glutamate receptor antagonist CNQX, but the agonist pharmacology is quite different from mammalian receptors. We are currently cloning excitatory receptors from Helisoma, and from another pond snail, Biomphalaria glabrata, an intermediate host for the parasite causing schistosomiasis in humans, based on sequence homology withLymnaea. Cloned receptors will be expressed in Xenopus oocytes to examine their characteristics for comparison to native receptors and receptors from other species.
The receptors that respond to glutamate with inhibition are also of interest. Both metabotropic and ionotropic glutamate receptors mediating inhibition have been observed in pulmonate molluscs, but none have been cloned, and those in Helisoma and its close relatives are as yet unidentified in terms of pharmacology or mechanism. We are currently testing potential glutamate agonists and antagonists that have been shown to interact with other inhibitory receptors in invertebrates for their ability to mimic glutamate inhibition of burst firing. We are also testing potential inhibitors of signal transduction pathways to determine how inhibition occurs. The goal is to develop a pharmacological and signal transduction profile of the receptor(s) to determine whether they are similar to any know invertebrate or vertebrate receptors. From this information a strategy will be developed for cloning these receptors and other proteins involved in inhibition.
Future interests include the further characterization of the role glutamate and its receptors play in the neuronal network controlling feeding behavior in the snails, particularly in Biomphalaria, for which little physiological information is available.
Mammalian glutamate receptors expressed in Xenopus oocytes
The ionotropic glutamate receptors consist of three subtypes of receptors, AMPA receptors, kainate receptors and NMDA receptors. All mediate fast excitatory neurotransmission in mammalian neurons by allowing cells to become depolarized. Of the three types, NMDA receptors are the most complex in terms of the normal functions they are involved in (development of the nervous system, learning and memory), the pathological conditions they affect (epilepsy, brain damage due to stroke, neurodegenerative disorders), and the number of binding sites they contain for modulators. My lab is interested in understanding what types of compounds (drugs or natural modulators) bind to NMDA receptors to affect receptor activity. We are also interested in understanding how the drugs bind (i.e., what the mechanism is).
To study how drugs affect NMDA receptor activity, different combinations of rat brain NMDA receptor subunits are expressed in Xenopus oocytes by injecting the oocytes with RNA derived from the cDNA clones. The oocytes make the receptor proteins and assemble them in their appropriate places in the oocyte membrane. When the receptors are activated by glutamate (or the selective agonist, NMDA) and the coagonist glycine, the ion channel contained in the receptors opens and ion movements can be measured. By varying the concentrations of drugs and neurotransmitters, potencies can be determined and the mechanism by which the drugs act can be examined. Some of the drugs we have investigated recently are the anticonvulsant felbamate, and the antipsychotics chlorpromazine and clozapine. See student projects for more details on these projects.