Jerzy W. Mozrzymas
Kinetics and modulation of synaptic receptors: how it looks like in the time scale of synaptic transmission ?
The time course of excitatory and inhibitory synaptic currents plays a key role in determining the integration of synaptic signals in neurons. The kinetic shape of these currents depends on the amount and time course of neurotransmitter released from the nerve terminal and the kinetics of the postsynaptic receptors. In the central nervous system synaptic agonist (e.g. GABA and glutamate) is known to reach millimolar concentrations and is cleared out from the synaptic cleft within hundreds of microseconds. Notably, the synaptic currents usually outlast by far the time duration of agonist transient implying that intrinsic kinetic properties of receptors play a major role in determining the time course of synaptic currents. Moreover, extremely brief presence of agonist within synaptic cleft indicates that the postsynaptic receptors are activated in conditions of extreme non-equilibrium. In this presentation I shall concentrate our attention on the inhibitory synaptic transmission for which GABA is the major neurotransmitter in the adult brain. Synaptic currents can be routinely measured in the whole-cell mode of the patch-clamp technique. However, the analysis of the IPSCs is insufficient to characterize the kinetics of postsynaptic receptors. The description of kinetic properties of ligand-gated channels requires construction of dose-response relationships for current responses elicited by exogenous agonist applications. However, the perfusion of agonist should be sufficiently fast to mimic the synaptic transient of the neurotransmitter. Using the ultrafast, piezoelectric driven perfusion system a complete exchange of agonist can be achieved within 100 μs largely fulfilling thus this requirement. The analysis of current responses to ultrafast GABA applications allowed characterizing the kinetics of these receptors with time resolution adequate to the synaptic events. The application of this technique made it possible also to describe the mechanism of pharmacological modulation of GABAA receptors by several clinically relevant compounds such as benzodiazepines, phenothiazines, barbiturates as well as by other modulators such as e.g. pH, Zn ions. The native GABAA receptors are known to be extremely heterogeneous. GABAA receptors composed of different subunits show different kinetic and pharmacological properties. For this reason, the expression of recombinant receptors with a strictly defined subunit composition in cell lines provided the means to further characterize the kinetics and pharmacology of the GABAA receptors in strict relation to its tetriary structure.