Time-resolved imaging of Ca(2+)-dependent aequorin luminescence of microdomains and QEDs in synaptic preterminals

Abstract

Localized elevation of intracellular free calcium [Ca2+]i concentration serves as the trigger for a wide variety of physiological processes, e.g., neurotransmitter release at most chemical synapses (1-3). The details of the mechanisms that regulate these processes are still unresolved (3-6), but they must involve precise temporal sequences of molecular events initiated by a transient localized elevation of Ca2+ concentration (i.e., a Ca2+ microdomain [3,7-15]). A microdomain is defined as an autonomous compartment of minimal spatio-temporal volume within which a signaled process can occur (8, 10, 12). A quantum emission domain (QED) is a quantal signal element (3, 16, 17). The concept of a QED was first applied to Ca2+ signaling at the synaptic preterminal (3, 4) and for large-diameter mitotic cells (16, 17). The concept of Ca2+ microdomains was tested by labeling preterminals of squid giant synapses with low-sensitivity aequorin (a photoprotein that emits a photon upon binding Ca2+ [18, 19]). That work confirmed earlier modeling efforts (10, 16) and showed that, upon depolarization, the [Ca2+]i profile reaches 200-300 microM within the microdomains, and that these [Ca2+]i profiles are composed of groups of short-lived 0.5 microns diameter QEDs. In those records, obtained with 2:1 interlacing devices operating at the RS-170 standard, QEDs appeared as striped dots or chevrons rather than as solid dots, indicating that a QED lasted less than 16.6 ms (one video field), and thus establishing the need for higher sampling rates to better characterize the QED.(ABSTRACT TRUNCATED AT 250 WORDS)