Two-photon scanning photochemical microscopy: mapping ligand-gated ion channel distributions.
The locations and densities of ionotropic membrane receptors, which are responsible for receiving synaptic transmission throughout the nervous system, are of prime importance in understanding the function of neural circuits. It is shown that the highly localized liberation of "caged" neurotransmitters by two-photon absorption-mediated photoactivation can be used in conjunction with recording the induced whole-cell current to determine the distribution of ligand-gated ion channels. The technique is potentially sensitive enough to detect individual channels with diffraction-limited spatial resolution. Images of the distribution of nicotinic acetylcholine receptors on cultured BC3H1 cells were obtained using a photoactivatable precursor of the nicotinic agonist carbamoylcholine.
Denk
W
W
AT&T Bell Laboratories, Murray Hill, NJ 07974.
D000818:Animals; D002217:Carbachol / Q000031:analogs & derivatives*; D002460:Cell Line; D005456:Fluorescent Dyes; D005463:Fluoroacetates; D015640:Ion Channel Gating; D007473:Ion Channels / Q000502:physiology* / Q000648:ultrastructure; D007834:Lasers; D008564:Membrane Potentials; D051379:Mice; D008853:Microscopy / Q000379:methods*; D009578:Nitrobenzenes; D017785:Photons; D011978:Receptors, Nicotinic / Q000032:analysis*; D012235:Rhodamines; D013997:Time Factors
D005456:Fluorescent Dyes; D005463:Fluoroacetates; D007473:Ion Channels; C088047:N-((alpha-carboxy)-2-nitrobenzyl)carbamoylcholine trifluoroacetate; D009578:Nitrobenzenes; D011978:Receptors, Nicotinic; D012235:Rhodamines; D002217:Carbachol; C029773:rhodamine B