Facilitation at single synapses probed with optical quantal analysis

Abstract

Many synapses can change their strength rapidly in a use-dependent manner, but the mechanisms of such short-term plasticity remain unknown. To understand these mechanisms, measurements of neurotransmitter release at single synapses are required. We probed transmitter release by imaging transient increases in [Ca(2+)] mediated by synaptic N-methyl-D-aspartate receptors (NMDARs) in individual dendritic spines of CA1 pyramidal neurons in rat brain slices, enabling quantal analysis at single synapses. We found that changes in release probability, produced by paired-pulse facilitation (PPF) or by manipulation of presynaptic adenosine receptors, were associated with changes in glutamate concentration in the synaptic cleft, indicating that single synapses can release a variable amount of glutamate per action potential. The relationship between release probability and response size is consistent with a binomial model of vesicle release with several (>5) independent release sites per active zone, suggesting that multivesicular release contributes to facilitation at these synapses.

The optimal height of the synaptic cleft

“…The synaptic active zone radius was set at 70 nm, and the number of available AMPA receptors varied from 50 to 120, in accordance with the detailed experimental description of the common glutamatergic synapses in area CA1 of the hippocampus (32)(33)(34). Similarly, the number of glutamate molecules released into the cleft was varied from the baseline value of 3,000 (35) to 8,000, to reflect the possibility of multivesicular release at this synaptic type (36). Finally, the lateral size of the synaptic apposition was varied 5-fold (between 200 and 1,000 nm), to reflect the high variability, including age-dependence, of the overall synaptic dimensions (10)(11)(12)19).…”

Quantal Size Is Independent of the Release Probability at Hippocampal Excitatory Synapses

“…It is generally agreed that an increase in the P r occurs during facilitation; however, it is still debated whether the increase in P r is solely responsible for the facilitation or whether postsynaptic modifications also take place. Several studies reported an exclusive presynaptic alteration (Gulyas et al, 1993;Stevens and Wang, 1995;Dobrunz and Stevens, 1997;Hanse and Gustafsson, 2001;Silver et al, 2003;Chen et al, 2004;Lawrence et al, 2004), but evidence has also been published supporting significant postsynaptic modifications, including an increased quantal size after multivesicular release (Oertner et al, 2002;Conti and Lisman, 2003) or removing the polyamine block from certain postsynaptic glutamate receptors (Rozov and Burnashev, 1999). To some extent, the reason for the discrepancy is likely attributable to differences in experimental approaches, such as the preparations, the age of the animals, and the identity of the synapses examined.…”

“…This led to the formulation of the one-vesicle hypothesis (Korn et al, 1994): either no vesicle or only one vesicle is released at one release site after the arrival of a presynaptic action potential. Several recent studies tested the one-release site, one-vesicle hypothesis at mammalian central synapses and came to various conclusions (Auger et al, 1998;Wadiche and Jahr, 2001;Oertner et al, 2002;Schneggenburger et al, 2002;Conti and Lisman, 2003) (but see Gulyas et al, 1993;Stevens and Wang, 1995;Dobrunz and Stevens, 1997;Hanse and Gustafsson, 2001;Silver et al, 2003;Lawrence et al, 2004). As mentioned above, the discrepancy may be the consequence of technical differences, most notably distinctions in the way presynaptic axons were stimulated.…”

“…For example, paired recordings with subsequent light microscopic (LM) and electron microscopic (EM) analysis of the number of synaptic contacts have been successful approaches to obtain independent estimates of N (Korn et al, 1982;Gulyas et al, 1993;Buhl et al, 1997;Silver et al, 2003). Alternatively, two-photon imaging of calcium transients in single spines (Oertner et al, 2002;Nimchinsky et al, 2004) and imaging of the destaining rate of FM1-43 fluorescence (Murthy et al, 1997) have been used to monitor P r independently of electrophysiological recordings. It is widely accepted that short-term plasticity is accompanied by changes in P r (for review, see Thomson, 2000;Zucker and Regehr, 2002); however, whether alterations in P r (purely presynaptic) fully account for such plasticity (Gulyas et al, 1993;Paulsen and Heggelund, 1994;Stevens and Wang, 1995;Hanse and Gustafsson, 2001;Silver et al, 2003;Lawrence et al, 2004) or whether postsynaptic alterations also occur (Otis et al, 1996;Rozov and Burnashev, 1999;Oleskevich et al, 2000;Oertner et al, 2002;Harrison and Jahr, 2003) is still a question of active debate.…”

“…Alternatively, two-photon imaging of calcium transients in single spines (Oertner et al, 2002;Nimchinsky et al, 2004) and imaging of the destaining rate of FM1-43 fluorescence (Murthy et al, 1997) have been used to monitor P r independently of electrophysiological recordings. It is widely accepted that short-term plasticity is accompanied by changes in P r (for review, see Thomson, 2000;Zucker and Regehr, 2002); however, whether alterations in P r (purely presynaptic) fully account for such plasticity (Gulyas et al, 1993;Paulsen and Heggelund, 1994;Stevens and Wang, 1995;Hanse and Gustafsson, 2001;Silver et al, 2003;Lawrence et al, 2004) or whether postsynaptic alterations also occur (Otis et al, 1996;Rozov and Burnashev, 1999;Oleskevich et al, 2000;Oertner et al, 2002;Harrison and Jahr, 2003) is still a question of active debate. To determine the quantal parameters and the locus of alterations during short-term facilitation at cortical glutamatergic synapses, we took advantage of the robust facilitation of synapses made by hippocampal CA1 pyramidal cells (PCs) onto specific subtypes of interneurons (Ali and Thomson, 1998;Scanziani et al, 1998;Losonczy et al, 2002).…”