A dynamin 1-, dynamin 3- and clathrin-independent pathway of synaptic vesicle recycling mediated by bulk endocytosis

Abstract

The exocytosis of synaptic vesicles (SVs) elicited by potent stimulation is rapidly compensated by bulk endocytosis of SV membranes leading to large endocytic vacuoles (‘bulk’ endosomes). Subsequently, these vacuoles disappear in parallel with the reappearance of new SVs. We have used synapses of dynamin 1 and 3 double knock-out neurons, where clathrin-mediated endocytosis (CME) is dramatically impaired, to gain insight into the poorly understood mechanisms underlying this process. Massive formation of bulk endosomes was not defective, but rather enhanced, in the absence of dynamin 1 and 3. The subsequent conversion of bulk endosomes into SVs was not accompanied by the accumulation of clathrin coated buds on their surface and this process proceeded even after further clathrin knock-down, suggesting its independence of clathrin. These findings support the existence of a pathway for SV reformation that bypasses the requirement for clathrin and dynamin 1/3 and that operates during intense synaptic activity.DOI: http://dx.doi.org/10.7554/eLife.01621.001

Tissue-specific dynamin-1 deletion at the calyx of Held decreases short-term depression through a mechanism distinct from vesicle resupply

“…The fast phase of Cm decay (at 2 s) in cKO tended to be faster (decreased to 88.0 ± 5.2% of its peak, n = 5) than in control (to 94.4 ± 3.1%, n = 7; P = 0.33). These data agree well with our previous work showing a pronounced impairment in the second Cm decay phase after strong but not after mild stimulation (28), as well as impaired clathrin-coated endocytosis under the electron microscope in dynamin-1 KO terminals (26,28,67). Remaining dynamin-2 and -3 may be sufficient to support low levels of endocytosis and transmission in the absence of dynamin-1.…”

Dynamin 2 regulates biphasic insulin secretion and plasma glucose homeostasis

“…A similar scenario has been reported at rodent nerve terminals lacking dynamin 1 (36,75), in which dynamin 3 partially supports vesicle recycling (38,39). It is unclear whether the dynamin-independent endocytosis at some nonsecretory cells (45,71) and nerve terminals (i.e., bulk endocytosis) (40,72,76) also occurs in dynamin 2 KO β cells. Future study using triple dynamin KO cells will shed new light on this question.…”

“…The meandering tubules in KO β cells were approximately 20 to 80 nm in diameter and up to hundreds of nanometers in length. These structures are reminiscent of endocytic defects in central synapses lacking dynamin 1 (and dynamin 3) (36,38,40) but more complicated than those in fibroblasts lacking dynamin (45). A unique feature of these endocytic intermediates is that each tubule was capped by a CCP, suggesting a severe defect in the membrane fission reactions of CME.…”

“…For electron tomography, tissue sections (200-250 nm) were imaged in a TECNAI TF20 transmission electron microscope (FEI) operated at 200 kV (Yale Center for Cellular and Molecular Imaging, Yale University) as described previously (40). Contours of membranes were traced using IMOD software manually.…”

“…Dynamin 1 is highly expressed in the nervous system, dynamin 2 is ubiquitous in all tissues, and dynamin 3 is found primarily in neuronal tissues and at low levels in testis (35,36). At nerve terminals, dynamin 1 and 3 contribute to synaptic vesicle reformation and neurotransmission (36)(37)(38)(39)(40); synapses lacking dynamin 1 (and dynamin 3) have fewer vesicles and abundant clathrin-coated profiles (CCPs) accumulated on the PM (36,38,40). In nonneuronal cells, dynamin isoforms appear cell-type specific and may play distinct roles in multiple endocytosis pathways (32,35,41).…”