In the vertebrate retina, presynaptic terminals of photoreceptors and bipolar cells form ribbon synapses and release neurotransmitter continuously. Endocytic machinery in the ribbon synapse is likely to differ from that in conventional synapses because of the much higher rate of synaptic vesicle recycling. However, protein components of the ribbon synapse identified so far are quite similar to those of the conventional synapses. Recently we identified amphiphysin I splice variants, termed amphiphysin Ir, that are transcribed specifically in retina from the authentic amphiphysin I gene [Y. Terada et al. (2002) FEBS Lett., 519, 185-190]. Amphiphysin I is a nerve terminal-enriched protein, and involved in synaptic vesicle endocytosis as heterodimer with amphiphysin II, an isoform of amphiphysin I. We report here that the retina-specific amphiphysin Ir is expressed exclusively in the ribbon synapse and not in conventional synapses. This is the first endocytosis-related, ribbon synapse-specific protein identified in the retina. By immunoprecipitation and double-immunolabelling, amphiphysin Ir was shown to be associated not only with amphiphysin II, but also with dynamin, clathrin and alpha-adaptin that are involved in synaptic vesicle recycling. The results suggest that endocytosis of the synaptic vesicle membrane in retinal ribbon synapses proceeds through a pathway similar to the one that is used in conventional synapses, although amphiphysin Ir is substituted for amphiphysin I. Amphiphysin Ir may play an essential role in the avid endocytic activity of ribbon synapses by associating with yet unknown protein partner(s) through its large insertional domain, which is absent from the conventional amphiphysin I.