Neuroscience Graduate Program at UCSF
Structure, Function and Regulation of Neuronal Nicotinic Acetylcholine Receptors
Our laboratory is interested in the role that nicotinic receptors (nAChRs) play in synaptic transmission. NAChRs are ionotropic receptors gated by the transmitter acetylcholine (ACh) and also the exogenous ligand nicotine. They underlie rapid synaptic transmission in the peripheral nervous system. The diversity of nAChR expression in the CNS and the widespread effects of nicotinic agonists on behavior imply that nAChRs are involved in a host of functions, including attention, learning and memory, and goal-directed behavior.
To enhance our understanding of nicotinic synaptic mechanisms we have studied transmission at a highly accessible cholinergic synapse in the peripheral nervous system: the giant calyciform synapse in the chick ciliary ganglion. This synapse consists of a large calyx terminating on a dendrite-free postsynaptic neuron. At late embryonic stages, the synapse is well-developed and allows the postsynaptic neurons to follow presynaptic action potentials at high frequencies. The post-synaptic neuron is electrically compact, and whole cell recordings can be made both from it and also from the presynaptic terminal. NAChRs are found both postsynaptically, where they underlie rapid synaptic transmission, and presynaptically, where their function is not known.
In the ciliary ganglion we have used imaging techniques and confocal microscopy to examine the distribution of AChRs on the surface of ciliary neurons, and neurophysiological techniques to study the role that distinct classes of nAChRs play in synaptic transmission in the ganglion. We have found that ACh activates at least two different AChRs: those containing a 7 subunits (a7-nAChRs) and those containing a3 subunits (a3*-nAChRs, the “*” indicates that other subunits are present in addition to a3). We have also shown that the distribution of these nAChRs is distinct: a3*-nAChRs feature prominently at synaptic sites, which are present both on the soma and on somatic spines, while both a7-nAChRs and a3*-nAChRs are found diffusely on the spines . The fact that extrasynaptic a7-nAChRs are activated by nerve-released ACh has led to the proposal that they are activated by ectopic release - release not occurring from active zones [Coggan et al. (2005) Science 309:446-451]. We are doing experiments to test this proposal and also to examine the role that ACh spillover plays in transmission at this synapse.
In a study published in 2009 we found that a3*-nAChRs did not contribute significantly to responses of the postsynaptic cell to individual quanta of ACh, perhaps because a3*-nAChRs at synaptic sites have a low probability of opening. Most ACh quanta simultaneously activate a7-nAChRs and a3*-nAChRs; since a7-nAChRs are largely confined to spines, most detectable release events must occur there, where both nAChR types are distributed diffusely. In a study published in 2011, we found that spillover of ACh is a prominent feature of transmission at this synapse, and we also found that the two nAChR classes do not “report” ACh spillover equally well; a3*-nAChRs readily respond to ACh that spills over from release sites, while a7-nAChRs do not. Indeed, our evidence suggests that a7-nAChRs buffer ACh and regulate its availability to a3*-nAChRs, mimicking the effect that extracellular ACh binding proteins have in other systems.
Recently, we found that ACh spillover activates nAChRs on glial cells in the ganglion (satellite cells) and produces very slowly rising and decaying currents that are indicative of paracrine or “volume” transmission. We are performing experiments to characterize this volume transmission, and we have applied for funding to extend these studies into cholinergic synapses in the mammalian brain (optogenetic mice), where volume transmission has been proposed to be an important signaling pathway for nicotinic receptors.
Dourado M, Sargent PB (2002) Properties of nicotinic receptors underlying Renshaw cell excitation by -motor neurons in neonatal rat spinal cord. J. Neurophysiol. 87: 3117-3125
Nguyen D, Sargent PB (2002) Synaptic vesicle recycling at two classes of release sites in giant nerve terminals of the embryonic chicken ciliary ganglion. J. Comp. Neurol. 448: 128-137.
Rogers M, Sargent PB (2003) Rapid activation of presynaptic nicotinic acetylcholine receptors by nerve-released transmitter. Eur J Neurosci 18: 2946-2956
Sargent PB, Saviane C, Nielsen TA, DiGregorio DA, Silver RA (2005) Rapid vesicular release, quantal variability, and spillover contribute to the precision and reliability of transmission at a glomerular synapse. J Neurosci 25:8173-8187.
Sargent PB (2009) Nicotinic receptors concentrated in the subsynaptic membrane do not contribute significantly to synaptic currents at an embryonic synapse in the chicken ciliary ganglion. J Neurosci 29: 3749-3759.
Stanchev D, Sargent PB (2011) α7-Containing and non-α7-containing nicotinic receptors respond differently to spillover of acetylcholine. J Neurosci 31: 14920-14930.