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Defective function of GABA-containing synaptic vesicles in mice lacking the AP-3B clathrin adaptor.

Nakatsu F, Okada M, Mori F, Kumazawa N, Iwasa H, Zhu G, Kasagi Y, Kamiya H, Harada A, Nishimura K, Takeuchi A, Miyazaki T, Watanabe M, Yuasa S, Manabe T, Wakabayashi K, Kaneko S, Saito T, Ohno H - J. Cell Biol. (2004)

Bottom Line: Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved.This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway.Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles.

View Article: PubMed Central - PubMed

Affiliation: RIKEN Research Center for Allergy and Immunology, Kanagawa 230-0045, Japan.

ABSTRACT
AP-3 is a member of the adaptor protein (AP) complex family that regulates the vesicular transport of cargo proteins in the secretory and endocytic pathways. There are two isoforms of AP-3: the ubiquitously expressed AP-3A and the neuron-specific AP-3B. Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved. To address this question, we generated mice lacking mu3B, a subunit of AP-3B. mu3B-/- mice suffered from spontaneous epileptic seizures. Morphological abnormalities were observed at synapses in these mice. Biochemical studies demonstrated the impairment of gamma-aminobutyric acid (GABA) release because of, at least in part, the reduction of vesicular GABA transporter in mu3B-/- mice. This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway. Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles. This work adds a new aspect to the pathogenesis of epilepsy.

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Enhanced synaptic potentiation through reduced GABAergic synaptic inhibition in μ3B−/−ΔNeo mice. (A–D) The average time course of the slope of synaptic responses in μ3B−/−ΔNeo mice and their littermate wild-type mice. Initial EPSP slopes were normalized in each experiment to the average slope value of the baseline (−30–0 min). The potentiation ratio was calculated by dividing the average slope value from 50 to 60 min by that of the baseline. Afferent fibers were tetanized at time 0 at: (A) 100 Hz for 1 s in the presence of PTX in wild-type (open circles; n = 18) and μ3B−/−ΔNeo (closed circles; n = 19) mice; (B) 100 Hz for 1 s in the absence of PTX in wild-type (open circles; n = 16) and μ3B−/−ΔNeo (closed circles; n = 13) mice; (C) 100 Hz for 200 ms in the presence of PTX in wild-type (open circles; n = 10) and μ3B−/−ΔNeo (closed circles; n = 10) mice; and (D) 100 Hz for 200 ms in the absence of PTX in wild-type (open circles; n = 14) and μ3B−/−ΔNeo (closed circles; n = 15) mice.
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fig5: Enhanced synaptic potentiation through reduced GABAergic synaptic inhibition in μ3B−/−ΔNeo mice. (A–D) The average time course of the slope of synaptic responses in μ3B−/−ΔNeo mice and their littermate wild-type mice. Initial EPSP slopes were normalized in each experiment to the average slope value of the baseline (−30–0 min). The potentiation ratio was calculated by dividing the average slope value from 50 to 60 min by that of the baseline. Afferent fibers were tetanized at time 0 at: (A) 100 Hz for 1 s in the presence of PTX in wild-type (open circles; n = 18) and μ3B−/−ΔNeo (closed circles; n = 19) mice; (B) 100 Hz for 1 s in the absence of PTX in wild-type (open circles; n = 16) and μ3B−/−ΔNeo (closed circles; n = 13) mice; (C) 100 Hz for 200 ms in the presence of PTX in wild-type (open circles; n = 10) and μ3B−/−ΔNeo (closed circles; n = 10) mice; and (D) 100 Hz for 200 ms in the absence of PTX in wild-type (open circles; n = 14) and μ3B−/−ΔNeo (closed circles; n = 15) mice.

Mentions: It is well established that the threshold for the induction of long-term potentiation (LTP) of excitatory synaptic transmission in the hippocampal CA1 region is regulated by GABAA receptor-mediated inhibitory synaptic inputs that are activated by afferent fiber stimulation for LTP induction (Wigstrom and Gustafsson, 1983): disinhibition by the blockade of GABAA receptor facilitates LTP induction. To test whether GABAergic synaptic inhibition is impaired in μ3B−/−ΔNeo mice, we examined the effect of picrotoxin (PTX; 100 μM), a GABAA receptor antagonist, on LTP induction. LTP induced by standard conditioning (100 Hz for 1 s) in μ3B−/−ΔNeo mice was intact either in the presence (P > 0.05; Fig. 5 A) or in the absence (P > 0.05; Fig. 5 B) of PTX. However, when weak conditioning (100 Hz for 200 ms) was applied in the absence of PTX (Fig. 5 D), LTP was not induced in wild-type mice (99.3 ± 1.4% of baseline), whereas stable potentiation was induced in μ3B−/−ΔNeo mice (117.7 ± 1.7% of baseline; P < 0.05). This difference disappeared when PTX was present (P > 0.05; Fig. 5 C), indicating that the phenotype observed in Fig. 5 D was dependent on GABAA receptor-mediated synaptic transmission. Thus, it is conceivable that when a weaker tetanus is used, the influence of inhibition is relatively stronger and LTP induction is suppressed in wild-type mice, whereas LTP is induced in μ3B−/−ΔNeo mice because the inhibition is weaker. These results suggest impaired GABAergic synaptic transmission in μ3B−/−ΔNeo mice, and are consistent with the reduced GABA release from presynaptic terminals in μ3B−/−ΔNeo mice.


Defective function of GABA-containing synaptic vesicles in mice lacking the AP-3B clathrin adaptor.

Nakatsu F, Okada M, Mori F, Kumazawa N, Iwasa H, Zhu G, Kasagi Y, Kamiya H, Harada A, Nishimura K, Takeuchi A, Miyazaki T, Watanabe M, Yuasa S, Manabe T, Wakabayashi K, Kaneko S, Saito T, Ohno H - J. Cell Biol. (2004)

Enhanced synaptic potentiation through reduced GABAergic synaptic inhibition in μ3B−/−ΔNeo mice. (A–D) The average time course of the slope of synaptic responses in μ3B−/−ΔNeo mice and their littermate wild-type mice. Initial EPSP slopes were normalized in each experiment to the average slope value of the baseline (−30–0 min). The potentiation ratio was calculated by dividing the average slope value from 50 to 60 min by that of the baseline. Afferent fibers were tetanized at time 0 at: (A) 100 Hz for 1 s in the presence of PTX in wild-type (open circles; n = 18) and μ3B−/−ΔNeo (closed circles; n = 19) mice; (B) 100 Hz for 1 s in the absence of PTX in wild-type (open circles; n = 16) and μ3B−/−ΔNeo (closed circles; n = 13) mice; (C) 100 Hz for 200 ms in the presence of PTX in wild-type (open circles; n = 10) and μ3B−/−ΔNeo (closed circles; n = 10) mice; and (D) 100 Hz for 200 ms in the absence of PTX in wild-type (open circles; n = 14) and μ3B−/−ΔNeo (closed circles; n = 15) mice.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2172536&req=5

fig5: Enhanced synaptic potentiation through reduced GABAergic synaptic inhibition in μ3B−/−ΔNeo mice. (A–D) The average time course of the slope of synaptic responses in μ3B−/−ΔNeo mice and their littermate wild-type mice. Initial EPSP slopes were normalized in each experiment to the average slope value of the baseline (−30–0 min). The potentiation ratio was calculated by dividing the average slope value from 50 to 60 min by that of the baseline. Afferent fibers were tetanized at time 0 at: (A) 100 Hz for 1 s in the presence of PTX in wild-type (open circles; n = 18) and μ3B−/−ΔNeo (closed circles; n = 19) mice; (B) 100 Hz for 1 s in the absence of PTX in wild-type (open circles; n = 16) and μ3B−/−ΔNeo (closed circles; n = 13) mice; (C) 100 Hz for 200 ms in the presence of PTX in wild-type (open circles; n = 10) and μ3B−/−ΔNeo (closed circles; n = 10) mice; and (D) 100 Hz for 200 ms in the absence of PTX in wild-type (open circles; n = 14) and μ3B−/−ΔNeo (closed circles; n = 15) mice.
Mentions: It is well established that the threshold for the induction of long-term potentiation (LTP) of excitatory synaptic transmission in the hippocampal CA1 region is regulated by GABAA receptor-mediated inhibitory synaptic inputs that are activated by afferent fiber stimulation for LTP induction (Wigstrom and Gustafsson, 1983): disinhibition by the blockade of GABAA receptor facilitates LTP induction. To test whether GABAergic synaptic inhibition is impaired in μ3B−/−ΔNeo mice, we examined the effect of picrotoxin (PTX; 100 μM), a GABAA receptor antagonist, on LTP induction. LTP induced by standard conditioning (100 Hz for 1 s) in μ3B−/−ΔNeo mice was intact either in the presence (P > 0.05; Fig. 5 A) or in the absence (P > 0.05; Fig. 5 B) of PTX. However, when weak conditioning (100 Hz for 200 ms) was applied in the absence of PTX (Fig. 5 D), LTP was not induced in wild-type mice (99.3 ± 1.4% of baseline), whereas stable potentiation was induced in μ3B−/−ΔNeo mice (117.7 ± 1.7% of baseline; P < 0.05). This difference disappeared when PTX was present (P > 0.05; Fig. 5 C), indicating that the phenotype observed in Fig. 5 D was dependent on GABAA receptor-mediated synaptic transmission. Thus, it is conceivable that when a weaker tetanus is used, the influence of inhibition is relatively stronger and LTP induction is suppressed in wild-type mice, whereas LTP is induced in μ3B−/−ΔNeo mice because the inhibition is weaker. These results suggest impaired GABAergic synaptic transmission in μ3B−/−ΔNeo mice, and are consistent with the reduced GABA release from presynaptic terminals in μ3B−/−ΔNeo mice.

Bottom Line: Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved.This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway.Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles.

View Article: PubMed Central - PubMed

Affiliation: RIKEN Research Center for Allergy and Immunology, Kanagawa 230-0045, Japan.

ABSTRACT
AP-3 is a member of the adaptor protein (AP) complex family that regulates the vesicular transport of cargo proteins in the secretory and endocytic pathways. There are two isoforms of AP-3: the ubiquitously expressed AP-3A and the neuron-specific AP-3B. Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved. To address this question, we generated mice lacking mu3B, a subunit of AP-3B. mu3B-/- mice suffered from spontaneous epileptic seizures. Morphological abnormalities were observed at synapses in these mice. Biochemical studies demonstrated the impairment of gamma-aminobutyric acid (GABA) release because of, at least in part, the reduction of vesicular GABA transporter in mu3B-/- mice. This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway. Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles. This work adds a new aspect to the pathogenesis of epilepsy.

Show MeSH
Related in: MedlinePlus