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Active zone protein Bassoon co-localizes with presynaptic calcium channel, modifies channel function, and recovers from aging related loss by exercise.

Nishimune H, Numata T, Chen J, Aoki Y, Wang Y, Starr MP, Mori Y, Stanford JA - PLoS ONE (2012)

Bottom Line: However, presynaptic Bassoon level was significantly attenuated in aged rat NMJs, which suggests an attenuation of VDCC function due to a lack of this interaction between VDCC and Bassoon.Importantly, the decreased Bassoon level in aged NMJs was ameliorated by isometric strength training of muscles for two months.However, Bassoon level in aged NMJs can be improved by muscle exercise.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Cell Biology, University of Kansas Medical School, Kansas City, Kansas, United States of America. hnishimune@kumc.edu

ABSTRACT
The P/Q-type voltage-dependent calcium channels (VDCCs) are essential for synaptic transmission at adult mammalian neuromuscular junctions (NMJs); however, the subsynaptic location of VDCCs relative to active zones in rodent NMJs, and the functional modification of VDCCs by the interaction with active zone protein Bassoon remain unknown. Here, we show that P/Q-type VDCCs distribute in a punctate pattern within the NMJ presynaptic terminals and align in three dimensions with Bassoon. This distribution pattern of P/Q-type VDCCs and Bassoon in NMJs is consistent with our previous study demonstrating the binding of VDCCs and Bassoon. In addition, we now show that the interaction between P/Q-type VDCCs and Bassoon significantly suppressed the inactivation property of P/Q-type VDCCs, suggesting that the Ca(2+) influx may be augmented by Bassoon for efficient synaptic transmission at NMJs. However, presynaptic Bassoon level was significantly attenuated in aged rat NMJs, which suggests an attenuation of VDCC function due to a lack of this interaction between VDCC and Bassoon. Importantly, the decreased Bassoon level in aged NMJs was ameliorated by isometric strength training of muscles for two months. The training increased Bassoon immunoreactivity in NMJs without affecting synapse size. These results demonstrated that the P/Q-type VDCCs preferentially accumulate at NMJ active zones and play essential role in synaptic transmission in conjunction with the active zone protein Bassoon. This molecular mechanism becomes impaired by aging, which suggests altered synaptic function in aged NMJs. However, Bassoon level in aged NMJs can be improved by muscle exercise.

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Inactivation properties of the P/Q-type VDCC were suppressed by Bassoon.(A) Left, inactivation of P/Q-type VDCC (Cav2.1) currents in BHK cells stably expressing VDCCs and transfected with an expression vector pBassoon-IRES2-GFP (Bsn, red) or an empty pIRES2-GFP vector (control, black). The peak amplitudes were normalized for Ba2+ currents elicited by 2-s pulses to 0 mV from a holding potential of –100 mV. Right graph shows inactivation curves for P/Q-type VDCCs with or without Bassoon. The half-inactivation potential was significantly higher in Bassoon expressing cells (–40.0±1.7 mV) compared to controls (–46.1±0.6 mV). (B) I-V relationships of P/Q-type VDCC showed no difference between with or without Bassoon. Left, representative traces are Ba2+ currents of Cav2.1 with or without Bassoon by applying test pluses from –100 mV (holding potential) to –50 mV up to 40 mV in 10 mV increments. Right graph shows current density-voltage (I-V) relationships. (C) The activation property of P/Q-type VDCCs in the presence of Bassoon exhibited a depolarization shift. Left, effects of Bassoon on activation of Cav2.1 currents elicited in BHK cells. Tail currents were elicited by repolarization to –60 mV after 5-ms test pulse from –50 to 55 mV with 5 mV increments. Right, activation curves were determined using these tail currents with or without Bassoon. The half-activation potential was significantly higher in Bassoon expressing cells (–0.7±1.1 mV) compared to controls (–4.0±1.1 mV). (D) Activation kinetics of P/Q-type VDCC currents. Left, tail currents were evoked by 5 ms depolarization from the holding potential (–100 mV) to 0 mV. Right graph shows the activation time constants (τ activation) with or without Bassoon. The activation time constant (τ) increased significantly in the presence of Bassoon at membrane potentials higher than 0 mV. Recordings from eight independent cells were averaged and the mean ± SEM are shown. Asterisks indicate significant difference compared to the controls, *p<0.05, **p<0.01.
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pone-0038029-g002: Inactivation properties of the P/Q-type VDCC were suppressed by Bassoon.(A) Left, inactivation of P/Q-type VDCC (Cav2.1) currents in BHK cells stably expressing VDCCs and transfected with an expression vector pBassoon-IRES2-GFP (Bsn, red) or an empty pIRES2-GFP vector (control, black). The peak amplitudes were normalized for Ba2+ currents elicited by 2-s pulses to 0 mV from a holding potential of –100 mV. Right graph shows inactivation curves for P/Q-type VDCCs with or without Bassoon. The half-inactivation potential was significantly higher in Bassoon expressing cells (–40.0±1.7 mV) compared to controls (–46.1±0.6 mV). (B) I-V relationships of P/Q-type VDCC showed no difference between with or without Bassoon. Left, representative traces are Ba2+ currents of Cav2.1 with or without Bassoon by applying test pluses from –100 mV (holding potential) to –50 mV up to 40 mV in 10 mV increments. Right graph shows current density-voltage (I-V) relationships. (C) The activation property of P/Q-type VDCCs in the presence of Bassoon exhibited a depolarization shift. Left, effects of Bassoon on activation of Cav2.1 currents elicited in BHK cells. Tail currents were elicited by repolarization to –60 mV after 5-ms test pulse from –50 to 55 mV with 5 mV increments. Right, activation curves were determined using these tail currents with or without Bassoon. The half-activation potential was significantly higher in Bassoon expressing cells (–0.7±1.1 mV) compared to controls (–4.0±1.1 mV). (D) Activation kinetics of P/Q-type VDCC currents. Left, tail currents were evoked by 5 ms depolarization from the holding potential (–100 mV) to 0 mV. Right graph shows the activation time constants (τ activation) with or without Bassoon. The activation time constant (τ) increased significantly in the presence of Bassoon at membrane potentials higher than 0 mV. Recordings from eight independent cells were averaged and the mean ± SEM are shown. Asterisks indicate significant difference compared to the controls, *p<0.05, **p<0.01.

Mentions: The most prominent effect of Bassoon on VDCCs was observed on inactivation parameters. The inactivation property of P/Q-type VDCC was significantly suppressed by the co-expressed Bassoon (Fig. 2A, Table 1). The half-inactivation potential was shifted by 6.1 mV in Bassoon expressing cells (−40.0±1.7 mV) compared to controls (−46.1±0.6 mV). We observed a significant depolarizing shift in the voltage dependence of inactivation. This effect of Bassoon on P/Q-type VDCC is similar to the effect of Rim1 on VDCCs [17], [19].


Active zone protein Bassoon co-localizes with presynaptic calcium channel, modifies channel function, and recovers from aging related loss by exercise.

Nishimune H, Numata T, Chen J, Aoki Y, Wang Y, Starr MP, Mori Y, Stanford JA - PLoS ONE (2012)

Inactivation properties of the P/Q-type VDCC were suppressed by Bassoon.(A) Left, inactivation of P/Q-type VDCC (Cav2.1) currents in BHK cells stably expressing VDCCs and transfected with an expression vector pBassoon-IRES2-GFP (Bsn, red) or an empty pIRES2-GFP vector (control, black). The peak amplitudes were normalized for Ba2+ currents elicited by 2-s pulses to 0 mV from a holding potential of –100 mV. Right graph shows inactivation curves for P/Q-type VDCCs with or without Bassoon. The half-inactivation potential was significantly higher in Bassoon expressing cells (–40.0±1.7 mV) compared to controls (–46.1±0.6 mV). (B) I-V relationships of P/Q-type VDCC showed no difference between with or without Bassoon. Left, representative traces are Ba2+ currents of Cav2.1 with or without Bassoon by applying test pluses from –100 mV (holding potential) to –50 mV up to 40 mV in 10 mV increments. Right graph shows current density-voltage (I-V) relationships. (C) The activation property of P/Q-type VDCCs in the presence of Bassoon exhibited a depolarization shift. Left, effects of Bassoon on activation of Cav2.1 currents elicited in BHK cells. Tail currents were elicited by repolarization to –60 mV after 5-ms test pulse from –50 to 55 mV with 5 mV increments. Right, activation curves were determined using these tail currents with or without Bassoon. The half-activation potential was significantly higher in Bassoon expressing cells (–0.7±1.1 mV) compared to controls (–4.0±1.1 mV). (D) Activation kinetics of P/Q-type VDCC currents. Left, tail currents were evoked by 5 ms depolarization from the holding potential (–100 mV) to 0 mV. Right graph shows the activation time constants (τ activation) with or without Bassoon. The activation time constant (τ) increased significantly in the presence of Bassoon at membrane potentials higher than 0 mV. Recordings from eight independent cells were averaged and the mean ± SEM are shown. Asterisks indicate significant difference compared to the controls, *p<0.05, **p<0.01.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3368936&req=5

pone-0038029-g002: Inactivation properties of the P/Q-type VDCC were suppressed by Bassoon.(A) Left, inactivation of P/Q-type VDCC (Cav2.1) currents in BHK cells stably expressing VDCCs and transfected with an expression vector pBassoon-IRES2-GFP (Bsn, red) or an empty pIRES2-GFP vector (control, black). The peak amplitudes were normalized for Ba2+ currents elicited by 2-s pulses to 0 mV from a holding potential of –100 mV. Right graph shows inactivation curves for P/Q-type VDCCs with or without Bassoon. The half-inactivation potential was significantly higher in Bassoon expressing cells (–40.0±1.7 mV) compared to controls (–46.1±0.6 mV). (B) I-V relationships of P/Q-type VDCC showed no difference between with or without Bassoon. Left, representative traces are Ba2+ currents of Cav2.1 with or without Bassoon by applying test pluses from –100 mV (holding potential) to –50 mV up to 40 mV in 10 mV increments. Right graph shows current density-voltage (I-V) relationships. (C) The activation property of P/Q-type VDCCs in the presence of Bassoon exhibited a depolarization shift. Left, effects of Bassoon on activation of Cav2.1 currents elicited in BHK cells. Tail currents were elicited by repolarization to –60 mV after 5-ms test pulse from –50 to 55 mV with 5 mV increments. Right, activation curves were determined using these tail currents with or without Bassoon. The half-activation potential was significantly higher in Bassoon expressing cells (–0.7±1.1 mV) compared to controls (–4.0±1.1 mV). (D) Activation kinetics of P/Q-type VDCC currents. Left, tail currents were evoked by 5 ms depolarization from the holding potential (–100 mV) to 0 mV. Right graph shows the activation time constants (τ activation) with or without Bassoon. The activation time constant (τ) increased significantly in the presence of Bassoon at membrane potentials higher than 0 mV. Recordings from eight independent cells were averaged and the mean ± SEM are shown. Asterisks indicate significant difference compared to the controls, *p<0.05, **p<0.01.
Mentions: The most prominent effect of Bassoon on VDCCs was observed on inactivation parameters. The inactivation property of P/Q-type VDCC was significantly suppressed by the co-expressed Bassoon (Fig. 2A, Table 1). The half-inactivation potential was shifted by 6.1 mV in Bassoon expressing cells (−40.0±1.7 mV) compared to controls (−46.1±0.6 mV). We observed a significant depolarizing shift in the voltage dependence of inactivation. This effect of Bassoon on P/Q-type VDCC is similar to the effect of Rim1 on VDCCs [17], [19].

Bottom Line: However, presynaptic Bassoon level was significantly attenuated in aged rat NMJs, which suggests an attenuation of VDCC function due to a lack of this interaction between VDCC and Bassoon.Importantly, the decreased Bassoon level in aged NMJs was ameliorated by isometric strength training of muscles for two months.However, Bassoon level in aged NMJs can be improved by muscle exercise.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Cell Biology, University of Kansas Medical School, Kansas City, Kansas, United States of America. hnishimune@kumc.edu

ABSTRACT
The P/Q-type voltage-dependent calcium channels (VDCCs) are essential for synaptic transmission at adult mammalian neuromuscular junctions (NMJs); however, the subsynaptic location of VDCCs relative to active zones in rodent NMJs, and the functional modification of VDCCs by the interaction with active zone protein Bassoon remain unknown. Here, we show that P/Q-type VDCCs distribute in a punctate pattern within the NMJ presynaptic terminals and align in three dimensions with Bassoon. This distribution pattern of P/Q-type VDCCs and Bassoon in NMJs is consistent with our previous study demonstrating the binding of VDCCs and Bassoon. In addition, we now show that the interaction between P/Q-type VDCCs and Bassoon significantly suppressed the inactivation property of P/Q-type VDCCs, suggesting that the Ca(2+) influx may be augmented by Bassoon for efficient synaptic transmission at NMJs. However, presynaptic Bassoon level was significantly attenuated in aged rat NMJs, which suggests an attenuation of VDCC function due to a lack of this interaction between VDCC and Bassoon. Importantly, the decreased Bassoon level in aged NMJs was ameliorated by isometric strength training of muscles for two months. The training increased Bassoon immunoreactivity in NMJs without affecting synapse size. These results demonstrated that the P/Q-type VDCCs preferentially accumulate at NMJ active zones and play essential role in synaptic transmission in conjunction with the active zone protein Bassoon. This molecular mechanism becomes impaired by aging, which suggests altered synaptic function in aged NMJs. However, Bassoon level in aged NMJs can be improved by muscle exercise.

Show MeSH
Related in: MedlinePlus