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Roles of KChIP1 in the regulation of GABA-mediated transmission and behavioral anxiety.

Xia K, Xiong H, Shin Y, Wang D, Deerinck T, Takahashi H, Ellisman MH, Lipton SA, Tong G, Descalzi G, Zhang D, Zhuo M, Zhang Z - Mol Brain (2010)

Bottom Line: Its physiological function, however, remains largely unknown.We report that KChIP1 is predominantly expressed at GABAergic synapses of a subset of parvalbumin-positive neurons in the brain.Forced expression of KChIP1 in cultured hippocampal neurons increased the frequency of miniature inhibitory postsynaptic currents (mIPSCs), reduced paired pulse facilitation of autaptic IPSCs, and decreases potassium current density.

View Article: PubMed Central - HTML - PubMed

Affiliation: State Key Laboratory of Medical Genetics, Central South University, Changsha, Hunan 410078, China.

ABSTRACT
K+ channel interacting protein 1 (KChIP1) is a neuronal calcium sensor (NCS) protein that interacts with multiple intracellular molecules. Its physiological function, however, remains largely unknown. We report that KChIP1 is predominantly expressed at GABAergic synapses of a subset of parvalbumin-positive neurons in the brain. Forced expression of KChIP1 in cultured hippocampal neurons increased the frequency of miniature inhibitory postsynaptic currents (mIPSCs), reduced paired pulse facilitation of autaptic IPSCs, and decreases potassium current density. Furthermore, genetic ablation of KChIP1 potentiated potassium current density in neurons and caused a robust enhancement of anxiety-like behavior in mice. Our study suggests that KChIP1 is a synaptic protein that regulates behavioral anxiety by modulating inhibitory synaptic transmission, and drugs that act on KChIP1 may help to treat patients with mood disorders including anxiety.

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Over-expression of KChIP1 facilitates GABA-mediated synaptic transmission in cultured hippocampal neurons. a: Expression of KChIP1-EGFP was detected by immunoblot analysis in neurons infected with KChIP1-EGFP (KChIP1) but not in neurons infected with EGFP alone. Detection of actin in the lysates with an anti-actin antibody is shown as a loading control. b: Representative recordings from EGFP+ (Control) and KChIP1-EGFP neurons (KChIP1) are shown. c: The paired pulse ratio (PPR) recorded in control cells was significantly larger than that seen in KChIP1-expressing cells (*P < 0.01). d, E: mIPSCs from autaptic inhibitory neurons. Representative mIPSC recordings are shown from EGFP+ (d) and KChIP1-EGFP+ neurons (e). f: Cumulative probability of amplitude and averaged mIPSCs (inset) from the same two cells as on f and e. g: The frequency of mIPSCs in cells expressing KChIP1 (0.88 ± 0.14 Hz) was more than twice that observed in control cells expressing EGFP alone (0.37 ± 0.13; *P < 0.05). h: There was no difference in the amplitude distribution of mIPSCs in these neurons (KChIP-EGFP: filled dots; EGFP alone: empty dots).
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Figure 3: Over-expression of KChIP1 facilitates GABA-mediated synaptic transmission in cultured hippocampal neurons. a: Expression of KChIP1-EGFP was detected by immunoblot analysis in neurons infected with KChIP1-EGFP (KChIP1) but not in neurons infected with EGFP alone. Detection of actin in the lysates with an anti-actin antibody is shown as a loading control. b: Representative recordings from EGFP+ (Control) and KChIP1-EGFP neurons (KChIP1) are shown. c: The paired pulse ratio (PPR) recorded in control cells was significantly larger than that seen in KChIP1-expressing cells (*P < 0.01). d, E: mIPSCs from autaptic inhibitory neurons. Representative mIPSC recordings are shown from EGFP+ (d) and KChIP1-EGFP+ neurons (e). f: Cumulative probability of amplitude and averaged mIPSCs (inset) from the same two cells as on f and e. g: The frequency of mIPSCs in cells expressing KChIP1 (0.88 ± 0.14 Hz) was more than twice that observed in control cells expressing EGFP alone (0.37 ± 0.13; *P < 0.05). h: There was no difference in the amplitude distribution of mIPSCs in these neurons (KChIP-EGFP: filled dots; EGFP alone: empty dots).

Mentions: We investigated the role of KChIP1 in modulating inhibitory synaptic transmission in cultured hippocampal neurons infected with recombinant Semliki Forest viral (SFV) particles encoding either a KChIP1-EGFP fusion protein or EGFP protein alone. Expression of KChIP1-EGFP was confirmed by immunoblotting with an anti-KChIP1 antibody (Fig. 3a). To assess its effect on physiological activity, we overexpressed KChIP1 in this neuronal cell type because only a subset of hippocampal neurons expressed endogenous KChIP1 and at relatively low levels. Using patch electrodes, we recorded autaptic GABA-mediated IPSCs from isolated EGFP-labeled neurons. IPSCs in these cultures were mediated exclusively by GABAA receptors that were blocked by the antagonists bicuculline or picrotoxinin [35]. Expression of KChIP1-EGFP had no significant effect on the decay time constants of autaptic GABA IPSCs (48.0 ± 6.7 ms, n = 7 for EGFP; 35.7 ± 3.0 ms, n = 8 for KChIP1-EGFP; P > 0.1 by Student's t-test). The peak amplitude of the IPSCs was 373 ± 108 pA (n = 8) in KChIP1-EGFP-positive neurons, while it was 233 ± 67.2 pA (n = 7) in EGFP-labeled neurons. Unlike the amplitude of GABA-mediated IPSCs, paired pulse facilitation is independent of postsynaptic receptor density or the number of synapses [36]. Here, we examined the effect of KChIP1 expression on this purely presynaptic phenomenon. Paired stimuli at intervals of 40 ms resulted in a 119 ± 12.8% increase in the amplitude of the second autaptic GABA IPSC in neurons expressing EGFP alone (n = 7). In contrast, for KChIP1-EGFP neurons, the paired pulse ratio was substantially reduced (74 ± 5.7%, n = 8; P < 0.01). This finding was consistent with the notion that KChIP1-EGFP expression increased the presynaptic release probability of GABA (Fig.3b and 3c). In agreement with the paired pulse facilitation results, the frequency of spontaneous autaptic miniature IPSCs (mIPSCs) in neurons expressing KChIP1-EGFP (0.88 ± 0.14 Hz, n = 12) was significantly higher than that seen in neurons expressing EGFP alone (0.37 ± 0.13 Hz, n = 10; P < 0.05) (Fig. 3d,e and 3g). However, there was no difference in the mean amplitude or amplitude distribution of mIPSCs in neurons expressing KChIP1-EGFP and control EGFP-positive neurons (Fig 3f and 3h). These results suggest that KChIP1 potentiates GABA-mediated inhibitory synaptic transmission via an increase in presynaptic GABA release.


Roles of KChIP1 in the regulation of GABA-mediated transmission and behavioral anxiety.

Xia K, Xiong H, Shin Y, Wang D, Deerinck T, Takahashi H, Ellisman MH, Lipton SA, Tong G, Descalzi G, Zhang D, Zhuo M, Zhang Z - Mol Brain (2010)

Over-expression of KChIP1 facilitates GABA-mediated synaptic transmission in cultured hippocampal neurons. a: Expression of KChIP1-EGFP was detected by immunoblot analysis in neurons infected with KChIP1-EGFP (KChIP1) but not in neurons infected with EGFP alone. Detection of actin in the lysates with an anti-actin antibody is shown as a loading control. b: Representative recordings from EGFP+ (Control) and KChIP1-EGFP neurons (KChIP1) are shown. c: The paired pulse ratio (PPR) recorded in control cells was significantly larger than that seen in KChIP1-expressing cells (*P < 0.01). d, E: mIPSCs from autaptic inhibitory neurons. Representative mIPSC recordings are shown from EGFP+ (d) and KChIP1-EGFP+ neurons (e). f: Cumulative probability of amplitude and averaged mIPSCs (inset) from the same two cells as on f and e. g: The frequency of mIPSCs in cells expressing KChIP1 (0.88 ± 0.14 Hz) was more than twice that observed in control cells expressing EGFP alone (0.37 ± 0.13; *P < 0.05). h: There was no difference in the amplitude distribution of mIPSCs in these neurons (KChIP-EGFP: filled dots; EGFP alone: empty dots).
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Figure 3: Over-expression of KChIP1 facilitates GABA-mediated synaptic transmission in cultured hippocampal neurons. a: Expression of KChIP1-EGFP was detected by immunoblot analysis in neurons infected with KChIP1-EGFP (KChIP1) but not in neurons infected with EGFP alone. Detection of actin in the lysates with an anti-actin antibody is shown as a loading control. b: Representative recordings from EGFP+ (Control) and KChIP1-EGFP neurons (KChIP1) are shown. c: The paired pulse ratio (PPR) recorded in control cells was significantly larger than that seen in KChIP1-expressing cells (*P < 0.01). d, E: mIPSCs from autaptic inhibitory neurons. Representative mIPSC recordings are shown from EGFP+ (d) and KChIP1-EGFP+ neurons (e). f: Cumulative probability of amplitude and averaged mIPSCs (inset) from the same two cells as on f and e. g: The frequency of mIPSCs in cells expressing KChIP1 (0.88 ± 0.14 Hz) was more than twice that observed in control cells expressing EGFP alone (0.37 ± 0.13; *P < 0.05). h: There was no difference in the amplitude distribution of mIPSCs in these neurons (KChIP-EGFP: filled dots; EGFP alone: empty dots).
Mentions: We investigated the role of KChIP1 in modulating inhibitory synaptic transmission in cultured hippocampal neurons infected with recombinant Semliki Forest viral (SFV) particles encoding either a KChIP1-EGFP fusion protein or EGFP protein alone. Expression of KChIP1-EGFP was confirmed by immunoblotting with an anti-KChIP1 antibody (Fig. 3a). To assess its effect on physiological activity, we overexpressed KChIP1 in this neuronal cell type because only a subset of hippocampal neurons expressed endogenous KChIP1 and at relatively low levels. Using patch electrodes, we recorded autaptic GABA-mediated IPSCs from isolated EGFP-labeled neurons. IPSCs in these cultures were mediated exclusively by GABAA receptors that were blocked by the antagonists bicuculline or picrotoxinin [35]. Expression of KChIP1-EGFP had no significant effect on the decay time constants of autaptic GABA IPSCs (48.0 ± 6.7 ms, n = 7 for EGFP; 35.7 ± 3.0 ms, n = 8 for KChIP1-EGFP; P > 0.1 by Student's t-test). The peak amplitude of the IPSCs was 373 ± 108 pA (n = 8) in KChIP1-EGFP-positive neurons, while it was 233 ± 67.2 pA (n = 7) in EGFP-labeled neurons. Unlike the amplitude of GABA-mediated IPSCs, paired pulse facilitation is independent of postsynaptic receptor density or the number of synapses [36]. Here, we examined the effect of KChIP1 expression on this purely presynaptic phenomenon. Paired stimuli at intervals of 40 ms resulted in a 119 ± 12.8% increase in the amplitude of the second autaptic GABA IPSC in neurons expressing EGFP alone (n = 7). In contrast, for KChIP1-EGFP neurons, the paired pulse ratio was substantially reduced (74 ± 5.7%, n = 8; P < 0.01). This finding was consistent with the notion that KChIP1-EGFP expression increased the presynaptic release probability of GABA (Fig.3b and 3c). In agreement with the paired pulse facilitation results, the frequency of spontaneous autaptic miniature IPSCs (mIPSCs) in neurons expressing KChIP1-EGFP (0.88 ± 0.14 Hz, n = 12) was significantly higher than that seen in neurons expressing EGFP alone (0.37 ± 0.13 Hz, n = 10; P < 0.05) (Fig. 3d,e and 3g). However, there was no difference in the mean amplitude or amplitude distribution of mIPSCs in neurons expressing KChIP1-EGFP and control EGFP-positive neurons (Fig 3f and 3h). These results suggest that KChIP1 potentiates GABA-mediated inhibitory synaptic transmission via an increase in presynaptic GABA release.

Bottom Line: Its physiological function, however, remains largely unknown.We report that KChIP1 is predominantly expressed at GABAergic synapses of a subset of parvalbumin-positive neurons in the brain.Forced expression of KChIP1 in cultured hippocampal neurons increased the frequency of miniature inhibitory postsynaptic currents (mIPSCs), reduced paired pulse facilitation of autaptic IPSCs, and decreases potassium current density.

View Article: PubMed Central - HTML - PubMed

Affiliation: State Key Laboratory of Medical Genetics, Central South University, Changsha, Hunan 410078, China.

ABSTRACT
K+ channel interacting protein 1 (KChIP1) is a neuronal calcium sensor (NCS) protein that interacts with multiple intracellular molecules. Its physiological function, however, remains largely unknown. We report that KChIP1 is predominantly expressed at GABAergic synapses of a subset of parvalbumin-positive neurons in the brain. Forced expression of KChIP1 in cultured hippocampal neurons increased the frequency of miniature inhibitory postsynaptic currents (mIPSCs), reduced paired pulse facilitation of autaptic IPSCs, and decreases potassium current density. Furthermore, genetic ablation of KChIP1 potentiated potassium current density in neurons and caused a robust enhancement of anxiety-like behavior in mice. Our study suggests that KChIP1 is a synaptic protein that regulates behavioral anxiety by modulating inhibitory synaptic transmission, and drugs that act on KChIP1 may help to treat patients with mood disorders including anxiety.

Show MeSH
Related in: MedlinePlus