Dynamics of Neuromuscular Transmission Reproduced by Calcium-Dependent and Reversible Serial Transitions in the Vesicle Fusion Complex

Abstract

Neuromuscular transmission, from spontaneous release to facilitation and depression, was accurately reproduced by a mechanistic kinetic model of sequential maturation transitions in the molecular fusion complex. The model incorporates three predictions. First, calcium-dependent forward transitions take vesicles from docked to preprimed to primed states, followed by fusion. Second, prepriming and priming are reversible. Third, fusion and recycling are unidirectional. The model was fed with experimental data from previous studies, whereas the backward (β) and recycling (ρ) rate constant values were fitted. Classical experiments were successfully reproduced with four transition states in the model when every forward (α) rate constant had the same value, and both backward rate constants were 50–100 times larger. Such disproportion originated an abruptly decreasing gradient of resting vesicles from docked to primed states. By contrast, a three-state version of the model failed to reproduce the dynamics of transmission by using the same set of parameters. Simulations predict the following: (1) Spontaneous release reflects primed to fusion spontaneous transitions. (2) Calcium elevations synchronize the series of forward transitions that lead to fusion. (3) Facilitation reflects a transient increase of priming following the calcium-dependent maturation transitions. (4) The calcium sensors that produce facilitation are those that evoke the transitions form docked to primed states. (5) Backward transitions and recycling restore the resting state. (6) Depression reflects backward transitions and slow recycling after intense release. Altogether, our results predict that fusion is produced by one calcium sensor, whereas the modulation of the number of vesicles that fuse depends on the calcium sensors that promote the early transition states. Such finely tuned kinetics offers a mechanism for collective non-linear transitional adaptations of a homogeneous vesicle pool to the ever-changing pattern of electrical activity in the neuromuscular junction.

Serotonin Promotes Vesicular Association and Fusion by Modifying Lipid Bilayers

“…We note that the vesicular fusion process in living cells is more complex and involves a series of priming steps regulated by the SM family and SNARE proteins before the actual release takes place. 18,19 For in vitro spontaneous fusion experiment used here, the vesicles that diffuse to the vicinity of the SLB can spontaneously bind to it, and then can either dissociate or irreversibly fuse with it (Fig. 2A).…”

“…17 The process is modelled by a series of calcium dependent maturation steps (docked pre-primed primed) before the actual fusion takes place where the pre-primed and primed states are reversible. 18,19 While the intracellular process of exocytosis involve the cellular machinery of various proteins 20 , it is useful to ask if the membrane itself can play any role in this process.…”

“…17 The process is modelled by a series of calcium dependent maturation steps (docked → pre-primed → primed) before the actual fusion takes place where the pre-primed and primed states are reversible. 18,19 While the intracellular process of exocytosis involve the cellular machinery of various proteins 20 , it is useful to ask if the membrane itself can play any role in this process. In our in vitro spontaneous fusion assay, we model this exocytosis process as reversible ‘docking’ and subsequent fusion of artificial lipid vesicles with supported lipid bilayers (SLBs).…”

Serotonin Promotes Vesicular Association and Fusion by Modifying Lipid Bilayers

“…We note that the vesicular fusion process in living cells is more complex and involves a series of priming steps regulated by the SM family and SNARE proteins before the actual release takes place. 14,40 For the in vitro spontaneous fusion experiment used here, the vesicles that diffuse to the vicinity of the SLB can spontaneously bind to it, and then can either dissociate or irreversibly fuse with it (Figure 2A). This process can be depicted by eq 5 (as described in Section 2.6):…”

A sequential two-step priming scheme reproduces diversity in synaptic strength and short-term plasticity

“…To more faithfully reproduce the multifaceted features of STP, different multiple-state and/or multiple-site schemes of transmitter release have been proposed ( 16 – 24 ), which include parallel schemes in which more than one kind of SV can bind to one or more kinds of release sites, and sequential schemes in which SVs migrate between different kinds of release sites or states of maturation.…”