James R Howe PhD
Professor of Pharmacology
We are interested in understanding how the structure of glutamate receptors determines their kinetic behavior. A combination of patch-clamp recording of macroscopic and single-channel currents and X-ray crystallography is employed to elucidate the major conformational changes that translate neurotransmitter binding into ion channel opening and receptor desensitization. Experimental and simulation studies are designed to determine the role of receptor kinetics in shaping synaptic transmission in the brain.
Extensive Research Description
Glutamate is the main excitatory neurotransmitter in the mammalian CNS and exerts many of its diverse effects through oligomeric receptors that are ligand-gated channels. Multiple families of glutamate-receptor (GluR) subunits exist, and each family contains several subunit genes. During the late 1990’s, most of our work focused on native channels. Using cerebellar granule cells as a convenient model system, we explored the expression and properties of GluRs in developing and mature neurons and investigated how the subunit composition of GluRs determines the functional properties of this important class of neuronal receptors. These studies revealed that developmental changes in kainate-type GluR subunit expression and processing are associated with changes in unitary channel properties (Smith et al., 1999). Other studies showed that granule cells begin to express multiple subtypes of AMPA-type GluR channels shortly before synaptogenesis and that some, but not all, of these subtypes are segregated to synapses in mature neurons (Ripellino et al., 1998; Smith et al., 2000). Both AMPA and kainate receptors in neurons in situ display multiple conductance levels. For AMPA receptors, the prevalence of sojourns in the different substates depends on the number of ligand molecules bound to the receptor (Smith and Howe, 2000). We also showed that alterations in the asynchrony of transmitter release and changes in receptor properties speed the kinetics of excitatory postsynaptic currents at mossy fiber-granule cell synapses as these synapses mature (Wall et al., 2002). Most of our recent work has focused on the kinetic behavior of recombinant glutamate receptors. A good deal about the structure of these receptors has emerged during the last several years, particularly for the ligand-binding domain, and we are trying to understand how major conformational changes impact receptor function. We provided the first functional evidence that AMPA receptors are dimers of dimers (Robert et al., 2001) and proposed new kinetic models that account for the structural data and differ significantly from previous kinetic schemes (Robert and Howe, 2003, Robert et al., 2005). We resolved a controversy regarding the effect of lurcher, a spontaneously arising mutation in a highly conserved motif in one of the transmembrane segments of AMPA receptors (Klein and Howe, 2004), and contributed to work showing that NMDA receptor kinetics influence the coincidence detection that underlies LTP (Popescu et al., 2004). We also demonstrated that an auxiliary protein, stargazin, has effects on the kinetics of single-channel currents that account for its slowing of the decay of excitatory postsynaptic currents (Tomita et al., 2005). In collaboration with Elias Lolis, we recently initiated crystallographic studies of the isolated GluR2 ligand-binding core.