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The active-zone protein Munc13 controls the use-dependence of presynaptic voltage-gated calcium channels.

Calloway N, Gouzer G, Xue M, Ryan TA - Elife (2015)

Bottom Line: Presynaptic calcium channel function is critical for converting electrical information into chemical communication but the molecules in the active zone that sculpt this function are poorly understood.We show that Munc13, an active-zone protein essential for exocytosis, also controls presynaptic voltage-gated calcium channel (VGCC) function dictating their behavior during various forms of activity.We demonstrate that in vitro Munc13 interacts with voltage-VGCCs via a pair of basic residues in Munc13's C2B domain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Weill Cornell Medical College, New York, United States.

ABSTRACT
Presynaptic calcium channel function is critical for converting electrical information into chemical communication but the molecules in the active zone that sculpt this function are poorly understood. We show that Munc13, an active-zone protein essential for exocytosis, also controls presynaptic voltage-gated calcium channel (VGCC) function dictating their behavior during various forms of activity. We demonstrate that in vitro Munc13 interacts with voltage-VGCCs via a pair of basic residues in Munc13's C2B domain. We show that elimination of this interaction by either removal of Munc13 or replacement of Munc13 with a Munc13 C2B mutant alters synaptic VGCC's response to and recovery from high-frequency action potential bursts and alters calcium influx from single action potential stimuli. These studies illustrate a novel form of synaptic modulation and show that Munc13 is poised to profoundly impact information transfer at nerve terminals by controlling both vesicle priming and the trigger for exocytosis.

No MeSH data available.


Related in: MedlinePlus

Munc13-KD prolongs VGCC inactivation on long-time scales.(A) Example of Ca2+ reactivation assay for WT and Munc13-KD neurons showing the difference in single AP responses before and after a 100 Hz 50 AP train showing that WT but not Munc13-KD cells have fully recovered from inactivation 500 ms after the end of the train. Inset shows AP following train on expanded time scale for more clarity. (B) Average inactivation of Ca2+ influx for WT and Munc13-KD neurons from the reactivation assay shown in A. Results are mean ± SEM. **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.07728.005
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fig3: Munc13-KD prolongs VGCC inactivation on long-time scales.(A) Example of Ca2+ reactivation assay for WT and Munc13-KD neurons showing the difference in single AP responses before and after a 100 Hz 50 AP train showing that WT but not Munc13-KD cells have fully recovered from inactivation 500 ms after the end of the train. Inset shows AP following train on expanded time scale for more clarity. (B) Average inactivation of Ca2+ influx for WT and Munc13-KD neurons from the reactivation assay shown in A. Results are mean ± SEM. **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.07728.005

Mentions: To investigate an underlying mechanism of Munc13's control of Ca2+ influx in presynaptic boutons that would be separate from changes in VGCC surface abundance, we first examined how the absence of Munc13 might influence the ability of VGCCs to recover from bursts of activity. Changes in Ca2+ influx after the burst would be consistent with alteration in VGCC kinetics of inactivation and/or recovery from protracted stimuli. To examine this issue, we compared single AP Ca2+ influx before and after a 0.5-s period of 100 Hz firing (Figure 3A). In WT synapses this level of activity did not impact single AP responses measured 0.5 s after the burst compared to before the burst. In contrast in the absence of Munc13, the single AP response 0.5 s after the burst was significantly lower (∼30%) than that measured before the burst (Figure 3A,B, Table 1). There are two possible explanations for this reduction: (1) in the absence of Munc13 there is increased VGCC inactivation following the 100 Hz stimulus and (2) in the absence of Munc13 VGCCs inactivate to the same extent as in WT but recover more slowly. At present, we do not have the means to separate these possibilities experimentally.10.7554/eLife.07728.005Figure 3.Munc13-KD prolongs VGCC inactivation on long-time scales.


The active-zone protein Munc13 controls the use-dependence of presynaptic voltage-gated calcium channels.

Calloway N, Gouzer G, Xue M, Ryan TA - Elife (2015)

Munc13-KD prolongs VGCC inactivation on long-time scales.(A) Example of Ca2+ reactivation assay for WT and Munc13-KD neurons showing the difference in single AP responses before and after a 100 Hz 50 AP train showing that WT but not Munc13-KD cells have fully recovered from inactivation 500 ms after the end of the train. Inset shows AP following train on expanded time scale for more clarity. (B) Average inactivation of Ca2+ influx for WT and Munc13-KD neurons from the reactivation assay shown in A. Results are mean ± SEM. **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.07728.005
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4525472&req=5

fig3: Munc13-KD prolongs VGCC inactivation on long-time scales.(A) Example of Ca2+ reactivation assay for WT and Munc13-KD neurons showing the difference in single AP responses before and after a 100 Hz 50 AP train showing that WT but not Munc13-KD cells have fully recovered from inactivation 500 ms after the end of the train. Inset shows AP following train on expanded time scale for more clarity. (B) Average inactivation of Ca2+ influx for WT and Munc13-KD neurons from the reactivation assay shown in A. Results are mean ± SEM. **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.07728.005
Mentions: To investigate an underlying mechanism of Munc13's control of Ca2+ influx in presynaptic boutons that would be separate from changes in VGCC surface abundance, we first examined how the absence of Munc13 might influence the ability of VGCCs to recover from bursts of activity. Changes in Ca2+ influx after the burst would be consistent with alteration in VGCC kinetics of inactivation and/or recovery from protracted stimuli. To examine this issue, we compared single AP Ca2+ influx before and after a 0.5-s period of 100 Hz firing (Figure 3A). In WT synapses this level of activity did not impact single AP responses measured 0.5 s after the burst compared to before the burst. In contrast in the absence of Munc13, the single AP response 0.5 s after the burst was significantly lower (∼30%) than that measured before the burst (Figure 3A,B, Table 1). There are two possible explanations for this reduction: (1) in the absence of Munc13 there is increased VGCC inactivation following the 100 Hz stimulus and (2) in the absence of Munc13 VGCCs inactivate to the same extent as in WT but recover more slowly. At present, we do not have the means to separate these possibilities experimentally.10.7554/eLife.07728.005Figure 3.Munc13-KD prolongs VGCC inactivation on long-time scales.

Bottom Line: Presynaptic calcium channel function is critical for converting electrical information into chemical communication but the molecules in the active zone that sculpt this function are poorly understood.We show that Munc13, an active-zone protein essential for exocytosis, also controls presynaptic voltage-gated calcium channel (VGCC) function dictating their behavior during various forms of activity.We demonstrate that in vitro Munc13 interacts with voltage-VGCCs via a pair of basic residues in Munc13's C2B domain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Weill Cornell Medical College, New York, United States.

ABSTRACT
Presynaptic calcium channel function is critical for converting electrical information into chemical communication but the molecules in the active zone that sculpt this function are poorly understood. We show that Munc13, an active-zone protein essential for exocytosis, also controls presynaptic voltage-gated calcium channel (VGCC) function dictating their behavior during various forms of activity. We demonstrate that in vitro Munc13 interacts with voltage-VGCCs via a pair of basic residues in Munc13's C2B domain. We show that elimination of this interaction by either removal of Munc13 or replacement of Munc13 with a Munc13 C2B mutant alters synaptic VGCC's response to and recovery from high-frequency action potential bursts and alters calcium influx from single action potential stimuli. These studies illustrate a novel form of synaptic modulation and show that Munc13 is poised to profoundly impact information transfer at nerve terminals by controlling both vesicle priming and the trigger for exocytosis.

No MeSH data available.


Related in: MedlinePlus