Fluorescent detection of Zn(2+)-rich vesicles with Zinquin: mechanism of action in lipid environments.
High concentrations of free Zn2+ ions are found in certain glutamatergic synaptic vesicles in the mammalian brain. These terminals can be visualized histochemically with quinoline sulfonamide compounds that form fluorescent complexes with Zn2+. The present study was undertaken to examine the interaction of the water-soluble quinoline sulfonamide probe, Zinquin (2-methyl-8-(toluene-p-sulfonamido)-6-quinolyloxyacetic acid) with the complex heterogeneous cellular environment. Experiments on rat hippocampal and neocortical slices gave indications that Zinquin in its free acid form was able to diffuse across the plasma and synaptic vesicle membranes. Further experiments were undertaken on unilamellar liposomes to study the interaction of Zinquin and its metal complexes in membranes. These experiments confirmed that Zinquin is able to diffuse across lipid bilayers. Steady-state and time-resolved fluorimetric studies showed that Zinquin in aqueous solution mainly forms a 1:2 (metal:ligand) complex with small amounts of a 1:1 complex. Formation of the 1:1 complex was favored by the presence of lipid, suggesting that it partitions into membranes. Evidence is presented that Zinquin can act as a Zn(2+)-ionophore, exchanging Zn2+ for two protons. The presence of a pH gradient across vesicles traps the Zn(2+)-probe complex within the vesicles. Zinquin is useful as a qualitative probe for detecting the presence of vesicular Zn2+; however, its tendency to partition into membranes and to serve as an ionophore should be borne in mind.
Snitsarev
V
V
Departmentsof Biological Sciences, University of Iowa, Iowa City 52242, USA.
Budde
T
T
Stricker
T P
TP
Cox
J M
JM
Krupa
D J
DJ
Geng
L
L
Kay
A R
AR
D000818:Animals; D001921:Brain / Q000166:cytology / Q000502:physiology*; D002462:Cell Membrane / Q000502:physiology; D004058:Diffusion; D005456:Fluorescent Dyes; D006624:Hippocampus / Q000166:cytology / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D008081:Liposomes; D008856:Microscopy, Fluorescence; D019579:Neocortex / Q000166:cytology / Q000502:physiology; D010713:Phosphatidylcholines; D015363:Quinolones / Q000493:pharmacokinetics; D051381:Rats; D020318:Rats, Long-Evans; D013050:Spectrometry, Fluorescence; D013572:Synaptic Vesicles / Q000502:physiology* / Q000648:ultrastructure; D014105:Tosyl Compounds / Q000493:pharmacokinetics; D015032:Zinc / Q000032:analysis* / Q000378:metabolism
D005456:Fluorescent Dyes; D008081:Liposomes; D010713:Phosphatidylcholines; D015363:Quinolones; D014105:Tosyl Compounds; C017251:1,2-oleoylphosphatidylcholine; D015032:Zinc; C084731:zinquin