Near-membrane [Ca2+] transients resolved using the
Ca2+ indicator FFP18
ELAINE F. ETTER*, AKWASI MINTAt, MARTIN POENIEt, AND FREDRIC S. FAY*
*Department of Physiology and Biomedical Imaging Group, University of Massachusetts Medical Center, 373 Plantation Street, Worcester, MA 01605;
tTEFLABS, 9503 Capitol View Drive, Austin, TX 78747; and *Department of Zoology, 141 Patterson Laboratories, University of Texas, Austin, TX 78712
Communicated by Joseph F. Hoffman, Yale University, New Haven, CT, December 22, 1995 (received for review August 28, 1995)
ABSTRACT: Ca2+-sensitive processes at cell membranes involved in contraction, secretion, and neurotransmitter release are activated in situ or in vitro by Ca2+ concentrations ([Ca2+]) 10-100 times higher than [Ca2+] measured during stimulation in intact cells. This paradox might be explained if the local [Ca2+] at the cell membrane is very different from that in the rest of the cell. Soluble Ca2+ indicators, which indicate spatially averaged cytoplasmic [Ca2+], cannot resolve these localized, near-membrane [Ca2+] signals. FFP18, the newest Ca2+ indicator designed to selectively monitor near-membrane [Ca2+], has a lower Ca2+ affinity and is more water soluble than previously used membrane-associating Ca2+ indicators. Images of the intracellular distribution of FFP18 show that >65% is located on or near the plasma membrane. [Cal] transients recorded using FFP18 during membrane depolarization induced Ca2+ influx show that near-membrane [Ca2+] rises faster and reaches micromolar levels at early times when the cytoplasmic [Ca2+], recorded using fura-2, has risen to only a few hundred nanomolar. High-speed series of digital images of [Ca+] show that near-membrane [Ca2+], reported by FFP18, rises within 20 msec, peaks at 50-100 msec, and then declines. [Ca+] reported by fura-2 rose slowly and continuously throughout the time images were acquired. The existence of these large, rapid increases in [Ca2+] directly beneath the surface membrane may explain how numerous Ca2+-sensitive membrane processes are activated at times when bulk cytoplasmic [Ca2+] changes are too small to activate them.
FFP-18 is now sold as Fura-2 NearMem (Near Membrane), the molecule remains unchanged.