Phosphorylation of cysteine string protein-α up-regulates the frequency of cholinergic waves via starburst amacrine cells.

During the first postnatal week in rodents, cholinergic retinal waves initiate in starburst amacrine cells (SACs), propagating to retinal ganglion cells (RGCs) and visual centers, essential for visual circuit refinement. By modulating exocytosis in SACs, dynamic changes in the protein kinase A (PKA) activity can regulate the spatiotemporal patterns of cholinergic waves. Previously, cysteine string protein-α (CSPα) is found to interact with the core exocytotic machinery by PKA-mediated phosphorylation at serine 10 (S10). However, whether PKA-mediated CSPα phosphorylation may regulate cholinergic waves via SACs remains unknown. Here, we examined how CSPα phosphorylation in SACs regulates cholinergic waves. First, we identified that CSPα1 is the major isoform in developing rat SACs and the inner plexiform layer during the first postnatal week. Using SAC-specific expression, we found that the CSPα1-PKA-phosphodeficient mutant (CSP-S10A) decreased wave frequency, but did not alter the wave spatial correlation compared to control, wild-type CSPα1 (CSP-WT), or two PKA-phosphomimetic mutants (CSP-S10D and CSP-S10E). These suggest that CSPα-S10 phosphodeficiency in SACs dampens the frequency of cholinergic waves. Moreover, the level of phospho-PKA substrates was significantly reduced in SACs overexpressing CSP-S10A compared to control or CSP-WT, suggesting that the dampened wave frequency is correlated with the decreased PKA activity. Further, compared to control or CSP-WT, CSP-S10A in SACs reduced the periodicity of wave-associated postsynaptic currents (PSCs) in neighboring RGCs, suggesting that these RGCs received the weakened synaptic inputs from SACs overexpressing CSP-S10A. Finally, CSP-S10A in SACs decreased the PSC amplitude and the slope to peak PSC compared to control or CSP-WT, suggesting that CSPα-S10 phosphodeficiency may dampen the speed of the SAC-RGC transmission. Thus, via PKA-mediated phosphorylation, CSPα in SACs may facilitate the SAC-RGC transmission, contributing to the robust frequency of cholinergic waves.

Syndesmosis Changes before and after Syndesmotic Screw Removal: A Retrospective Radiographic Study.

BACKGROUND AND OBJECTIVES: In patients with ankle fractures complicated by syndesmotic injuries, no consensus has been reached on the best method of syndesmosis fixation using syndesmotic screws. One previous study revealed no difference in the tibiofibular overlap between two groups with or without syndesmotic screw removal. Other studies have indicated that distal tibiofibular diastasis exists after the removal of syndesmotic screws. In this study, we aimed to confirm the effect of syndesmotic screw removal on diastasis occurrence. We further analyzed the risk factors that may contribute to the widening of the tibiofibular syndesmosis. MATERIALS AND METHODS: This retrospective study involved a review of the records of 63 patients with ankle fractures accompanied by syndesmosis injuries that required syndesmotic screw fixation. Anteroposterior radiographs were analyzed for each patient at various time points, from syndesmotic screw fixation to outpatient department follow-ups after screw removal. The changes in tibia-fibula overlap (OL), tibia-fibula clear space (CS), and medial clear space (MCS) were analyzed. Further analysis was performed to reveal potential factors that may have contributed to radiographic differences. RESULTS: Compared with the postoperation radiographs following syndesmotic screw fixation, OL decreased (2.0 mm) and CS increased (0.8 mm) in the anteroposterior radiographs from outpatient department follow-ups. No significant changes were noted in OL or CS after syndesmotic screw removal. However, OL decreased (1.8 mm) and CS increased (0.5 mm) before syndesmotic screw removal. No significant change in MCS occurred during the whole observation period. Linear regression analysis did not reveal any significant correlations between potentially related factors and radiographic changes. CONCLUSIONS: Marked diastasis had occurred at final follow-up. Notably, the diastasis occurred before rather than after screw removal. This implies that screw removal does not significantly influence the radiographic outcomes of rotational ankle fractures.

The Phosphoprotein Synapsin Ia Regulates the Kinetics of Dense-Core Vesicle Release.

Common fusion machinery mediates the Ca2+-dependent exocytosis of synaptic vesicles (SVs) and dense-core vesicles (DCVs). Previously, Synapsin Ia (Syn Ia) was found to localize to SVs, essential for mobilizing SVs to the plasma membrane through phosphorylation. However, whether (or how) the phosphoprotein Syn Ia plays a role in regulating DCV exocytosis remains unknown. To answer these questions, we measured the dynamics of DCV exocytosis by using single-vesicle amperometry in PC12 cells (derived from the pheochromocytoma of rats of unknown sex) overexpressing wild-type or phosphodeficient Syn Ia. We found that overexpression of phosphodeficient Syn Ia decreased the DCV secretion rate, specifically via residues previously shown to undergo calmodulin-dependent kinase (CaMK)-mediated phosphorylation (S9, S566, and S603). Moreover, the fusion pore kinetics during DCV exocytosis were found to be differentially regulated by Syn Ia and two phosphodeficient Syn Ia mutants (Syn Ia-S62A and Syn Ia-S9,566,603A). Kinetic analysis suggested that Syn Ia may regulate the closure and dilation of DCV fusion pores via these sites, implying the potential interactions of Syn Ia with certain DCV proteins involved in the regulation of fusion pore dynamics. Furthermore, we predicted the interaction of Syn Ia with several DCV proteins, including Synaptophysin (Syp) and soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins. By immunoprecipitation, we found that Syn Ia interacted with Syp via phosphorylation. Moreover, a proximity ligation assay (PLA) confirmed their phosphorylation-dependent, in situ interaction on DCVs. Together, these findings reveal a phosphorylation-mediated regulation of DCV exocytosis by Syn Ia.SIGNIFICANCE STATEMENT Although they exhibit distinct exocytosis dynamics upon stimulation, synaptic vesicles (SVs) and dense-core vesicles (DCVs) may undergo co-release in neurons and neuroendocrine cells through an undefined molecular mechanism. Synapsin Ia (Syn Ia) is known to recruit SVs to the plasma membrane via phosphorylation. Here, we examined whether Syn Ia also affects the dynamics of DCV exocytosis. We showed that Syn Ia regulates the DCV secretion rate and fusion pore kinetics during DCV exocytosis. Moreover, Syn Ia-mediated regulation of DCV exocytosis depends on phosphorylation. We further found that Syn Ia interacts with Synaptophysin (Syp) on DCVs in a phosphorylation-dependent manner. Thus, these results suggest that Syn Ia may regulate the release of DCVs via phosphorylation.

Presynaptic SNAP-25 regulates retinal waves and retinogeniculate projection via phosphorylation.

Patterned spontaneous activity periodically displays in developing retinas termed retinal waves, essential for visual circuit refinement. In neonatal rodents, retinal waves initiate in starburst amacrine cells (SACs), propagating across retinal ganglion cells (RGCs), further through visual centers. Although these waves are shown temporally synchronized with transiently high PKA activity, the downstream PKA target important for regulating the transmission from SACs remains unidentified. A t-SNARE, synaptosome-associated protein of 25 kDa (SNAP-25/SN25), serves as a PKA substrate, implying a potential role of SN25 in regulating retinal development. Here, we examined whether SN25 in SACs could regulate wave properties and retinogeniculate projection during development. In developing SACs, overexpression of wild-type SN25b, but not the PKA-phosphodeficient mutant (SN25b-T138A), decreased the frequency and spatial correlation of wave-associated calcium transients. Overexpressing SN25b, but not SN25b-T138A, in SACs dampened spontaneous, wave-associated, postsynaptic currents in RGCs and decreased the SAC release upon augmenting the cAMP-PKA signaling. These results suggest that SN25b overexpression may inhibit the strength of transmission from SACs via PKA-mediated phosphorylation at T138. Moreover, knockdown of endogenous SN25b increased the frequency of wave-associated calcium transients, supporting the role of SN25 in restraining wave periodicity. Finally, the eye-specific segregation of retinogeniculate projection was impaired by in vivo overexpression of SN25b, but not SN25b-T138A, in SACs. These results suggest that SN25 in developing SACs dampens the spatiotemporal properties of retinal waves and limits visual circuit refinement by phosphorylation at T138. Therefore, SN25 in SACs plays a profound role in regulating visual circuit refinement.

Characterization of information-based learning benefits with submovement dynamics and muscular rhythmicity.

For skill advancement, motor variability must be optimized based on target information during practice sessions. This study investigated structural changes in kinematic variability by characterizing submovement dynamics and muscular oscillations after practice with visuomotor tracking under different target conditions. Thirty-six participants were randomly assigned to one of three groups (simple, complex, and random). Each group practiced tracking visual targets with trajectories of varying complexity. The velocity trajectory of tracking was decomposed into 1) a primary contraction spectrally identical to the target rate and 2) an intermittent submovement profile. The learning benefits and submovement dynamics were conditional upon experimental manipulation of the target information. Only the simple and complex groups improved their skills with practice. The size of the submovements was most greatly reduced by practice with the least target information (simple > complex > random). Submovement complexity changed in parallel with learning benefits, with the most remarkable increase in practice under a moderate amount of target information (complex > simple > random). In the simple and complex protocols, skill improvements were associated with a significant decline in alpha (8-12 Hz) muscular oscillation but a potentiation of gamma (35-50 Hz) muscular oscillation. However, the random group showed no significant change in tracking skill or submovement dynamics, except that alpha muscular oscillation was reduced. In conclusion, submovement and gamma muscular oscillation are biological markers of learning benefits. Effective learning with an appropriate amount of target information reduces the size of submovements. In accordance with the challenge point hypothesis, changes in submovement complexity in response to target information had an inverted-U function, pertaining to an abundant trajectory-tuning strategy with target exactness.

Trajectory adjustments underlying task-specific intermittent force behaviors and muscular rhythms.

Force intermittency is one of the major causes of motor variability. Focusing on the dynamics of force intermittency, this study was undertaken to investigate how force trajectory is fine-tuned for static and dynamic force-tracking of a comparable physical load. Twenty-two healthy adults performed two unilateral resistance protocols (static force-tracking at 75% maximal effort and dynamic force-tracking in the range of 50%-100% maximal effort) using the left hand. The electromyographic activity and force profile of the designated hand were monitored. Gripping force was off-line decomposed into a primary movement spectrally identical to the target motion and a force intermittency profile containing numerous force pulses. The results showed that dynamic force-tracking exhibited greater intermittency amplitude and force pulse but a smaller amplitude ratio of primary movement to force intermittency than static force-tracking. Multi-scale entropy analysis revealed that force intermittency during dynamic force-tracking was more complex on a low time scale but more regular on a high time scale than that of static force-tracking. Together with task-dependent force intermittency properties, dynamic force-tracking exhibited a smaller 8-12 Hz muscular oscillation but a more potentiated muscular oscillation at 35-50 Hz than static force-tracking. In conclusion, force intermittency reflects differing trajectory controls for static and dynamic force-tracking. The target goal of dynamic tracking is achieved through trajectory adjustments that are more intricate and more frequent than those of static tracking, pertaining to differing organizations and functioning of muscular oscillations in the alpha and gamma bands.

Kinematic property of target motion conditions gaze behavior and eye-hand synergy during manual tracking.

This study investigated how frequency demand and motion feedback influenced composite ocular movements and eye-hand synergy during manual tracking. Fourteen volunteers conducted slow and fast force-tracking in which targets were displayed in either line-mode or wave-mode to guide manual tracking with target movement of direct position or velocity nature. The results showed that eye-hand synergy was a selective response of spatiotemporal coupling conditional on target rate and feedback mode. Slow and line-mode tracking exhibited stronger eye-hand coupling than fast and wave-mode tracking. Both eye movement and manual action led the target signal during fast-tracking, while the latency of ocular navigation during slow-tracking depended on the feedback mode. Slow-tracking resulted in more saccadic responses and larger pursuit gains than fast-tracking. Line-mode tracking led to larger pursuit gains but fewer and shorter gaze fixations than wave-mode tracking. During slow-tracking, incidences of saccade and gaze fixation fluctuated across a target cycle, peaking at velocity maximum and the maximal curvature of target displacement, respectively. For line-mode tracking, the incidence of smooth pursuit was phase-dependent, peaking at velocity maximum as well. Manual behavior of slow or line-mode tracking was better predicted by composite eye movements than that of fast or wave-mode tracking. In conclusion, manual tracking relied on versatile visual strategies to perceive target movements of different kinematic properties, which suggested a flexible coordinative control for the ocular and manual sensorimotor systems.

Eye-hand synergy and intermittent behaviors during target-directed tracking with visual and non-visual information.

Visual feedback and non-visual information play different roles in tracking of an external target. This study explored the respective roles of the visual and non-visual information in eleven healthy volunteers who coupled the manual cursor to a rhythmically moving target of 0.5 Hz under three sensorimotor conditions: eye-alone tracking (EA), eye-hand tracking with visual feedback of manual outputs (EH tracking), and the same tracking without such feedback (EHM tracking). Tracking error, kinematic variables, and movement intermittency (saccade and speed pulse) were contrasted among tracking conditions. The results showed that EHM tracking exhibited larger pursuit gain, less tracking error, and less movement intermittency for the ocular plant than EA tracking. With the vision of manual cursor, EH tracking achieved superior tracking congruency of the ocular and manual effectors with smaller movement intermittency than EHM tracking, except that the rate precision of manual action was similar for both types of tracking. The present study demonstrated that visibility of manual consequences altered mutual relationships between movement intermittency and tracking error. The speed pulse metrics of manual output were linked to ocular tracking error, and saccade events were time-locked to the positional error of manual tracking during EH tracking. In conclusion, peripheral non-visual information is critical to smooth pursuit characteristics and rate control of rhythmic manual tracking. Visual information adds to eye-hand synchrony, underlying improved amplitude control and elaborate error interpretation during oculo-manual tracking.

Organization of physiological tremors and coordination solutions to postural pointing on an uneven stance surface.

The study investigated the destabilization effect on multi-segment physiological tremors and coordinative control for a postural-suprapostural task under different stance conditions. Twenty volunteers executed postural pointing from a level surface and a seesaw balance board; meanwhile, physiological tremors of the whole postural system and fluctuation movements of fingertip/stance surface were recorded. In reference to level stance, seesaw stance led to much fewer tremor increments of the upper limb and less fluctuation movement of the fingertip than tremor increment of the lower limb and rolling movement of the stance surface. Tremor coupling between the adjacent segments organized differentially with stance surface. In reference to level stance, seesaw stance reinforced tremor coupling of the upper limb but enfeebled the coupling in the arm-lumbar and calf-foot complexes. Stance-related differences in physiological tremors could be explained by characteristic changes in the primary and secondary principal components (PC1 and PC2), with relatively high communality with segment tremors of the lower and upper limbs, respectively. Seesaw stance introduced a prominent 4-8Hz spectral peak in PC1 and potentiated 1-4Hz and 8-12Hz spectral peaks of PC2. Structural reorganization of physiological tremors with stance configuration suggests that seesaw stance involves distinct suprapostural and postural synergies for regulating degree of freedom in joint space.

Reorganization of multidigit physiological tremors after repetitive contractions of a single finger.

In light of the interplay among physiological finger tremors, this study was undertaken to investigate the transfer effect of fatigue on coordinative strategies of multiple fingers. Fourteen volunteers performed prolonged position tracking with a loaded middle finger while measures of neuromuscular function, including electromyographic activities of the extensor digitorum (ED)/flexor digitorum superficialis (FDS) and physiological tremors of the index, middle, ring, and little fingers, were monitored. The subjects exhibited inferior tracking congruence and an increase in ED activity at the end of the tracking. Fatigue spread was manifested in a remarkable increase in tremor across fingers, in association with enhanced involuntary tremor coupling among fingers that was topologically organized in relation to the distance of the digits from the middle finger. Principal component analysis suggested that an enhanced 8- to 12-Hz central rhythm contributed primarily to the tremor restructure following fatigue spread. The observed tremor reorganization validated the hypothesis that the effect of fatigue was not limited to the instructed finger and that fatigue functionally decreased independence of the digits. The spreading of fatigue weakens neural inputs that diverge to motor units acting on various digits because of fatigue-related enhancement of common drive at the supraspinal level.

Age effect on fatigue-induced limb acceleration as a consequence of high-level sustained submaximal contraction.

In reference to electromyographic measurement, the study was conducted to reassess differences in the behavior of fatigue-related neuromuscular function between young and elderly humans with limb acceleration (LA). Fourteen young and fourteen elderly subjects performed sustained index abduction at 75% of their maximal voluntary contractions (MVC) until task failure. Measures of neuromuscular function, including temporal/spectral features of muscle activity of the first dorsal interosseous (FDI) and LA of the index and hand, were monitored. The results showed a manifest fatigue-induced increase in LA of the index in the elderly group, but not in the young group. In contrast, only the young group developed a significant increase in amplitude of the electromyography (EMG) until task failure. Spectral analyses of LA in the index reflected marked age-dependent reorganization following muscle fatigue, with a greater reduction of relative spectral amplitude of LA in the range of 20-40 Hz, but a lesser reduction in coherence between EMG and LA in the elderly group. In line with fatigue-associated restructuring of LA, the mechanical coupling of the metacarpophalangeal joint was more severely undermined in the elderly group than in the young group. The present study manifested an age-related difference in the relative contributions of neural versus mechanical factors to muscle fatigue. Subsequent to a high-level sustained submaximal isometric contraction, a predominant mechanical failure of the musculotendon complex in the elderly was featured with LA, whereas EMG measurement characterized prevailing impairment of neuromuscular propagation in the young.

Exertion dependent alternations in force fluctuation and limb acceleration during sustained fatiguing contraction.

The study was conducted to contrast exertion-dependent changes in electromyography (EMG), force fluctuation (FF), and limb acceleration (LA) during isometric contraction to attain a versatile picture of muscle fatigue. Fifteen volunteers performed sustained index abduction at 25 and 75% maximal voluntary contractions (MVC) until failure at the tasks; meanwhile, changes in temporal/spectral features of force, muscle activity of the first dorsal interosseous (FDI), and acceleration of the index and hand were monitored. The results showed a manifest increase in all recorded signals for the 25% MVC paradigm, especially for LA, which demonstrated the largest increment in amplitude. In addition to progressive enhancement of the mechanical coupling of the metacarpophalangeal (MCP) joint, the 25% MVC paradigm added to EMG-FF and EMG-LA coherences (CohEMG-FF and CohEMG-LA) at 8-12 Hz and the shift of the spectral peak of the LA to higher frequencies. In contrast, the 75% MVC paradigm did not modulate significantly the spectral peak of LA. Also, CohEMG-FF, CohEMG-LA at 8-12 Hz, and the mechanical coupling of the MCP joint, were conversely undermined consequent to the high exertion paradigm. The present study suggests that LA was most susceptible to muscle fatigue following sustained contraction at a lower exertion level, and diverging alternations among various physiological signals ascribed to exertion-dependent contributions of central and peripheral origins to muscle fatigue.

Postural fluctuations during pointing from a unilateral or bilateral stance.

An experiment was conducted to compare the effects of bilateral and unilateral stance on postural fluctuations and intralimb coordination during active balance control. Fifteen participants stood bilaterally and unilaterally while conducting a pointing task with an outstretched arm. Excursion of center of foot pressure (CoP) and limb movements were recorded with a force plate and eight dual-axis accelerometers, respectively. Compared to bilateral stance, unilateral stance resulted in wider CoP trajectories and greater postural fluctuations, especially in the lower limbs. The limb-dependent postural fluctuations during unilateral stance were associated with an increased coupling between the upper limb segments and a decreased coupling between the segments of the stance leg. Unilateral stance further resulted in greater regularity and spectral changes in postural fluctuations of the trunk and lower limb due to increased central oscillations (8-15 Hz). The observed structural differences in postural fluctuations between unilateral and bilateral stance strongly suggested that the postural control system modulates joint stiffness in a stance-dependent manner. Probably, in unilateral stance, attentive control was shifted to the stance leg at the expense of increasing arm stiffness to reduce movement redundancy.