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

Abstract

Dynamin is a large GTPase with a crucial role in synaptic vesicle regeneration. Acute dynamin inhibition impairs neurotransmitter release, in agreement with the protein's established role in vesicle resupply. Here, using tissue-specific dynamin-1 knockout [conditional knockout (cKO)] mice at a fast central synapse that releases neurotransmitter at high rates, we report that dynamin-1 deletion unexpectedly leads to enhanced steady-state neurotransmission and consequently less synaptic depression during brief periods of high-frequency stimulation. These changes are also accompanied by increased transmission failures. Interestingly, synaptic vesicle resupply and several other synaptic properties remain intact, including basal neurotransmission, presynaptic Ca 2+ influx, initial release probability, and postsynaptic receptor saturation and desensitization. However, acute application of Latrunculin B, a reagent known to induce actin depolymerization and impair bulk and ultrafast endocytosis, has a stronger effect on steady-state depression in cKO than in control and brings the depression down to a control level. The slow phase of presynaptic capacitance decay following strong stimulation is impaired in cKO; the rapid capacitance changes immediately after strong depolarization are also different between control and cKO and sensitive to Latrunculin B. These data raise the possibility that, in addition to its established function in regenerating synaptic vesicles, the endocytosis protein dynamin-1 may have an impact on short-term synaptic depression. This role comes into play primarily during brief highfrequency stimulation.short-term plasticity | release site clearance | dynamin | bulk endocytosis | actin S ustained neurotransmission requires synaptic vesicle (SV) recycling. The reformation of functional SVs, in general, requires several seconds or even longer (1, 2) and is thus several orders of magnitude slower than vesicle release at active zones (AZs). Vesicle fusion can reach rates of up to a few thousand vesicles per second in many types of nerve terminals, such as ribbon synapses (3,000/s) (3), cerebellar basket cell terminals (5,000/s) (4), and the calyx of Held (6,000/s) (5-7). Because release sites are limited in number, synapses need to reuse them rapidly in succession during repetitive stimulation, and this requirement may be more restrictive than that of SV availability (8). AZs are composed of a meshwork of evolutionarily conserved scaffold proteins including RIM, Munc-13, RIM-BP, α-liprin, ELKS, and Ca 2+ channels (9-11), but their dynamic properties are poorly understood (10). During high-frequency release, vesicle membrane components (12) and soluble N-Ethylmaleimide sensitive factor (NSF) attachment protein receptor (SNARE) proteins that occupy the release sites need to be cleared rapidly so that new vesicles can dock and prime for new rounds of exocytosis. The recovery of release sites may become rate-limiting during high rates of transmitter release (8). However, direct experimental testing is chall...

Dynamin‐1 deletion enhances post‐tetanic potentiation and quantal size after tetanic stimulation at the calyx of Held

“…With this induction protocol, PTP is smaller at the mature CH (P16-20) than at the immature CH of pre-hearing animals, consistent with a developmental decrease of PTP under moderate induction (Korogod et al 2005). The basal EPSC amplitude in cKO (ß6 nA) was not significantly different from control (ß7 nA) (V h = −80 mV, without offline correction) (Mahapatra et al 2016). Interestingly, PTP is significantly higher in cKO (141.4 ± 6.6%, n = 11 neurons) than control (P = 0.007) ( Fig.…”

“…Direct presynaptic capacitance changes ( C m ) induced by a 50 ms depolarization did not reveal a significant difference in basal RRP and presynaptic Ca 2+ influx between cKO and control (Mahapatra et al . ), despite these results diverging from what is observed in conventional dynamin‐1 KO (presumably due to the healthier condition of cKO mice than conventional KOs; see Methods).…”

“…The basal EPSC amplitude in cKO (∼6 nA) was not significantly different from control (∼7 nA) ( V h = −80 mV, without offline correction) (Mahapatra et al . ). Interestingly, PTP is significantly higher in cKO (141.4 ± 6.6%, n = 11 neurons) than control ( P = 0.007) (Fig.…”

“…Dynamin‐1 deletion was confirmed in cKO mice ( Dnm1 fΔ Krox20 Cre/+ or Dnm1 f/f Krox20 Cre/+ ) (Mahapatra et al . ), and their littermates carrying no Krox20 Cre ( Dnm 1 f/Δ or Dnm1 f/f ) were used as control mice. The cKO mice had no outward phenotype compared to their littermate controls.…”

“…Dnm1 f/ Krox20 Cre/+ mice were further crossed with Dnm1 f/f to obtain conditional dynamin-1 KO (Dnm1 f/ Krox20 Cre/+ ) mice, in which Cre expression deletes the floxed Dnm1 gene selectively in Krox20 Cre -positive mice from embryonic day (E)9-10 (expression onset of Krox20 Cre ) onwards (Voiculescu et al 2000). Dynamin-1 deletion was confirmed in cKO mice (Dnm1 f Krox20 Cre/+ or Dnm1 f/f Krox20 Cre/+ ) (Mahapatra et al 2016), and their littermates carrying no Krox20 Cre (Dnm1 f/ or Dnm1 f/f ) were used as control mice. The cKO mice had no outward phenotype compared to their littermate controls.…”