Microdomains of High Calcium Concentration in a Presynaptic Terminal

Abstract

Increases in intracellular calcium concentration are required for the release of neurotransmitter from presynaptic terminals in all neurons. However, the mechanism by which calcium exerts its effect is not known. A low-sensitivity calcium-dependent photoprotein (n-aequorin-J) was injected into the presynaptic terminal of the giant squid synapse to selectively detect high calcium concentration microdomains. During transmitter release, light emission occurred at specific points or quantum emission domains that remained in the same place during protracted stimulation. Intracellular calcium concentration microdomains on the order of 200 to 300 micromolar occur against the cytoplasmic surface of the plasmalemma during transmitter secretion, supporting the view that the synaptic vesicular fusion responsible for transmitter release is triggered by the activation of a low-affinity calcium-binding site at the active zone.

Presynaptic plasticity: The regulation of Ca2+-dependent transmitter release

“…Therefore, a subceilular organization appears to exist in which Ca 2+ channels are very close to the synaptic vesicles at the active zone (see Zucker and Stockbridge, 1983;Simon and Llinas, 1985;Zucker and Fogelson, 1986;Smith and Augustine, 1989). This is supported by ultrastructural observations (see Dreyer, 1973;Heuser et al, 1979;Pumplin et al, 1981) and fluorescent monitoring of Ca 2+ (see Smith and Augustine, 1988;Swandulla et al, 1991;Llinas et al, 1992). Furthermore, the highest density of presynaptic Ca 2+ channels is located directly opposite to the highest density of postsynaptic receptors (Robitaille et al, 1990).…”

“…In this case plasticity of fast transmitter release may be entirely dependent on factors within this putative "Ca 2 + domain". Some evidence for such domains comes from the squid giant synapse (Llinas et al, 1992) and our preliminary data using confocal scanning microscopy suggest that some mammalian CNS synapses may also contain such restricted areas. Alternatively, local Ca2+-elevations may be obtained for longer periods without physical barriers if cytosolic Ca 2 + buffering is potent enough to absorb all Ca 2 + ions that diffuse from the submembrane area.…”

“…The selectivity of the process for Ca 2 + may in this case arise from the selectivity of the Ca 2 ÷ channels, rather than from the selectivity of the fusion machinery. Support for this idea can be found in the limited correlation between bulk intracellular [Ca 2 +]-elevations and release (Verhage et al, 1991b;Llinas et al, 1992), the poor effectivity of Ca 2 + released from photolabile chelators in the bulk cytosol (H6chner et al, 1989), the poor inhibition of release by slow-kinetics Ca 2 +-chelators (Adler et al, 1991) and the fact that divalent cations that can pass through Ca 2 + channels and accumulate inside nerve terminals, such as Ba 2 +, are also able to replace Ca 2 + in triggering transmitter release, but also with relatively low affinity (EDs0 ~ 0.5 mM).…”

“…However, it has been difficult to obtain unambiguous experimental evidence, mainly for practical reasons such as the poor accessibility of CNS nerve terminals to electrophysiological studies and Ca 2~ imaging. Studies on an unusually large nerve terminal, that of the squid giant axon, have revealed the occurrence of Ca -,+ "hot spots" associated with the tips of this terminal (Smith and Augustine, 1988;Swandulla et al, 1991;Llinas et al, 1992). We were able to obtain other, indirect evidence for the relevance of local elevations of [Ca]~ in CNS nerve terminals with the use of the mobile-carrier Ca ~' ~-ionophore ionomycin (Verhage et al, 1991b (Verhage et aL, 1991c) (Fig.…”

Complexity of calcium signaling in synaptic spines

“…Ca 2þ entry, particularly through NMDARs, which are localized on the synaptic face of the spine, will create a concentration gradient across the spine, with high concentrations, as high as hundreds of mM, near the mouth of the channel. (50) The amount of time [Ca 2þ ] i needs to be elevated to induce either LTP or LTD is a function of the probability of Ca 2þ ions encountering and activating sufficient numbers of specific target proteins. For uncaging experiments, this can take a long time given the relatively small number of free Ca 2þ ions (see below), particularly if these proteins are neither rapidly diffusing nor homogenously distributed.…”