Localization of the alpha(1F) calcium channel subunit in the rat retina.
OBJECTIVE
The molecular identity of the calcium channels that mediate glutamate release from photoreceptors is unknown. Mutations in the recently identified, retina-specific alpha(1F) calcium channel subunit cause incomplete X-linked congenital stationary night blindness (CSNB2), the phenotype of which is consistent with a defect in neurotransmission within the retina. The purpose of this study was to determine the cellular distribution of the alpha(1F) subunit in the retina.


METHODS
Antibodies were raised against a unique peptide from the human alpha(1F) sequence. Rat retina sections were labeled with affinity-purified alpha(1F) antibodies and the immunofluorescence analyzed by confocal microscopy. The alpha(1F) staining was compared with that obtained with a pan-alpha(1) antibody, used to reveal the distribution of known voltage-gated calcium channels in the retina. Some sections were double labeled for alpha(1F) and the photoreceptor synaptic ribbon marker, bassoon.


RESULTS
Staining of retina sections with anti-alpha(1F) resulted in strong punctate labeling in the outer plexiform layer (OPL) and weak punctate labeling in the inner plexiform layer (IPL), consistent with a synaptic localization. Staining was also observed in the outer nuclear layer. Within the OPL, alpha(1F) immunoreactivity was clustered in discrete, horseshoe-shaped patches, the shape and dimensions of which are characteristic of rod active zones. Similar structures were labeled with the pan-alpha(1) antibody. Localization of alpha(1F) immunoreactivity to rod active zones was confirmed by double labeling for bassoon, a component of photoreceptor synaptic ribbons.


CONCLUSIONS
The distribution of alpha(1F) immunoreactivity in the OPL suggests that calcium influx through alpha(1F) or alpha(1F)-like channels mediates glutamate release from rod photoreceptors.
Morgans
C W
CW
Division of Neuroscience, John Curtin School of Medical Research, Australian National University, Canberra, Australia. catherine.morgans@anu.edu.au
D000818:Animals; D015153:Blotting, Western; D015220:Calcium Channels / Q000378:metabolism*; D020746:Calcium Channels, L-Type; D005136:Eye Proteins / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D018613:Microscopy, Confocal; D009755:Night Blindness / Q000378:metabolism; D051381:Rats; D011921:Rats, Inbred WKY; D012160:Retina / Q000378:metabolism*; D012756:Sheep
C113330:CACNA1F protein, human; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D005136:Eye Proteins