Aberrant function and structure of retinal ribbon synapses in the absence of complexin 3 and complexin 4

Abstract

Complexins regulate the speed and Ca2+ sensitivity of SNARE-mediated synaptic vesicle fusion at conventional synapses. Two of the vertebrate complexins, Cplx3 and Cplx4, are specifically localized to retinal ribbon synapses. To test whether Cplx3 and Cplx4 contribute to the highly efficient transmitter release at ribbon synapses, we studied retina function and structure in Cplx3 and Cplx4 single- and double-knockout mice. Electroretinographic recordings from single and double mutants revealed a cooperative perturbing effect of Cplx3 and Cplx4 deletion on the b-wave amplitude, whereas most other detected effects in both plexiform synaptic layers were additive. Light and electron microscopic analyses uncovered a disorganized outer plexiform layer in the retinae of mice lacking Cplx3 and Cplx4, with a significant proportion of photoreceptor terminals containing spherical free-floating ribbons. These structural and functional aberrations were accompanied by behavioural deficits indicative of a vision deficit. Our results show that Cplx3 and Cplx4 are essential regulators of transmitter release at retinal ribbon synapses. Their loss leads to aberrant adjustment and fine-tuning of transmitter release at the photoreceptor ribbon synapse, alterations in transmission at bipolar cell terminals, changes in the temporal structure of synaptic processing in the inner plexiform layer of the retina and perturbed vision.

Munc13-Independent Vesicle Priming at Mouse Photoreceptor Ribbon Synapses

“…Molecular constituents of the photoreceptor ribbon synapses vary to some extent from those of conventional synapses and are likely best suited to fulfill these specific release requirements. They include the less prototypical SNAREs synaptobrevin-1 (Sherry et al, 2003) and syntaxin-3b (Morgans et al, 1996;Sherry et al, 2006;Curtis et al, 2008), and complexin-3 and -4 (Reim et al, 2009). Interestingly, the ERG phenotype reported in complexin-3/4 DKO mice (Reim et al, 2009) resembles the one observed in Munc13-2 KOs.…”

“…Electroretinographic (ERG) recordings were performed on five pairs of age-matched (8-to 12-week-old) male wild-type or Munc13-2 KO mice that were dark-adapted overnight before commencement of the measurements. Both scotopic and photopic responses were recorded and analyzed as previously described (Reim et al, 2009). Shortly, mice were anesthetized by intraperitoneal injections of xylazine (50 mg/kg) and ketamine (20 mg/kg).…”

“…From this filtered response, peak-to-peak amplitudes of OPs were measured. In addition, a time slice from the maximum of the a-wave to 200 ms was chosen for fast Fourier transformation (FFT) (Reim et al, 2009). In the FFT power spectrum, peak power and frequency at which peak power occurred were measured and plotted against light intensity.…”

Regulation of Exocytosis by Complexin

“…Systematic analyses of interactions between Cpx and SNARE complexes by employing both glutathione S-transferase (GST)-pulldowns and a yeast four-hybrid interaction assay showed that Cpx binds efficiently to the SNARE complexes that are composed of either Syntaxin 1 or Syntaxin 3 [17][18][19][20] . Cpx does not bind efficiently to the SNARE complexes that are composed of either Syntaxin 2 or Syntaxin 4 [18,20] .…”

“…Four Cpx isoforms (Cpx1-4) are expressed mainly in the nervous system such as the brain, spinal cord, and retina, where neurotransmitters are released in a synchronous manner ( Figure 2A) [13,16] . In particular, membranebound Cpx3 and Cpx4 are expressed in retinal ribbon synapses where strong synchronous neurotransmitter release is required [17] .…”