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GABAA receptor activity shapes the formation of inhibitory synapses between developing medium spiny neurons.

Arama J, Abitbol K, Goffin D, Fuchs C, Sihra TS, Thomson AM, Jovanovic JN - Front Cell Neurosci (2015)

Bottom Line: When activity of GABAARs was under chronic blockade between 4-7 DIV, the structural properties of these synapses remained unchanged.In contrast, chronic inhibition of GABAARs between 7-14 DIV led to reduction in size of α1- and α1/α2-postsynaptic clusters and a concomitant increase in number and size of α2-postsynaptic clusters.Thus, the main subtypes of GABAergic synapses formed by MSNs are regulated by GABAAR activity, but in opposite directions, and thus appear to be driven by different molecular mechanisms.

View Article: PubMed Central - PubMed

Affiliation: UCL School of Pharmacy, University College London London, UK.

ABSTRACT
Basal ganglia play an essential role in motor coordination and cognitive functions. The GABAergic medium spiny neurons (MSNs) account for ~95% of all the neurons in this brain region. Central to the normal functioning of MSNs is integration of synaptic activity arriving from the glutamatergic corticostriatal and thalamostriatal afferents, with synaptic inhibition mediated by local interneurons and MSN axon collaterals. In this study we have investigated how the specific types of GABAergic synapses between the MSNs develop over time, and how the activity of GABAA receptors (GABAARs) influences this development. Isolated embryonic (E17) MSNs form a homogenous population in vitro and display spontaneous synaptic activity and functional properties similar to their in vivo counterparts. In dual whole-cell recordings of synaptically connected pairs of MSNs, action potential (AP)-activated synaptic events were detected between 7 and 14 days in vitro (DIV), which coincided with the shift in GABAAR operation from depolarization to hyperpolarization, as detected indirectly by intracellular calcium imaging. In parallel, the predominant subtypes of inhibitory synapses, which innervate dendrites of MSNs and contain GABAAR α1 or α2 subunits, underwent distinct changes in the size of postsynaptic clusters, with α1 becoming smaller and α2 larger over time, while both the percentage and the size of mixed α1/α2-postsynaptic clusters were increased. When activity of GABAARs was under chronic blockade between 4-7 DIV, the structural properties of these synapses remained unchanged. In contrast, chronic inhibition of GABAARs between 7-14 DIV led to reduction in size of α1- and α1/α2-postsynaptic clusters and a concomitant increase in number and size of α2-postsynaptic clusters. Thus, the main subtypes of GABAergic synapses formed by MSNs are regulated by GABAAR activity, but in opposite directions, and thus appear to be driven by different molecular mechanisms.

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Related in: MedlinePlus

GABAA receptor activity has no influence on synapse formation between immature MSNs. (A,B) Immunolabeling of GABAA receptor α1- (i,ii; pink), α2- (iii,iv; green), α1/α2- (v,vi; white) subunit-containing clusters, and presynaptic GABAergic terminals (ii, iv, vi; red) along the primary dendrites of 7 DIV MSNs in the presence of vehicle control dimethy sulfoxide (DMSO) or Bic (25 μM), respectively. (i–vi) Scale bar = 5 μm. (C) The number of GABAergic terminals forming connections with primary dendrites of MSNs (n = 43 dendrites in DMSO/controls, n = 50 dendrites in Bic-treated cultures). (D–F) The number, percentage and size of synaptic α1-, α2- and α1/α2-clusters, respectively, along the primary dendrites of MSNs. The box plots display the median and IQRs of indicated synaptic parameters measured along the first 20 μm of primary dendrites (n = 49 dendrites in DMSO/controls, n = 46 dendrites in Bic-treated cultures) of total of n = 16 neurons from two independent experiments. Statistical analysis was performed using Mann Whitney test, *p < 0.05.
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Figure 8: GABAA receptor activity has no influence on synapse formation between immature MSNs. (A,B) Immunolabeling of GABAA receptor α1- (i,ii; pink), α2- (iii,iv; green), α1/α2- (v,vi; white) subunit-containing clusters, and presynaptic GABAergic terminals (ii, iv, vi; red) along the primary dendrites of 7 DIV MSNs in the presence of vehicle control dimethy sulfoxide (DMSO) or Bic (25 μM), respectively. (i–vi) Scale bar = 5 μm. (C) The number of GABAergic terminals forming connections with primary dendrites of MSNs (n = 43 dendrites in DMSO/controls, n = 50 dendrites in Bic-treated cultures). (D–F) The number, percentage and size of synaptic α1-, α2- and α1/α2-clusters, respectively, along the primary dendrites of MSNs. The box plots display the median and IQRs of indicated synaptic parameters measured along the first 20 μm of primary dendrites (n = 49 dendrites in DMSO/controls, n = 46 dendrites in Bic-treated cultures) of total of n = 16 neurons from two independent experiments. Statistical analysis was performed using Mann Whitney test, *p < 0.05.

Mentions: To establish whether depolarising activity of GABAA receptors has a regulatory role in synapse formation at the early stage of MSN development, and contrast this with the hyperpolarizing activity of GABAA receptors at the later stage, we have applied a chronic blockade of GABAA receptors using a competitive antagonist Bic (25 μM) in MSN cultures from 4–7 DIV or 7–14 DIV. We have first established that, at this concentration of Bic, the survival of MSNs was not impaired (data not shown). Using the immunocytochemistry approach and analysis described above, we subsequently characterized the formation of GABAergic synapses and the number and size of postsynaptic GABAA receptor clusters (Figures 8A,B).


GABAA receptor activity shapes the formation of inhibitory synapses between developing medium spiny neurons.

Arama J, Abitbol K, Goffin D, Fuchs C, Sihra TS, Thomson AM, Jovanovic JN - Front Cell Neurosci (2015)

GABAA receptor activity has no influence on synapse formation between immature MSNs. (A,B) Immunolabeling of GABAA receptor α1- (i,ii; pink), α2- (iii,iv; green), α1/α2- (v,vi; white) subunit-containing clusters, and presynaptic GABAergic terminals (ii, iv, vi; red) along the primary dendrites of 7 DIV MSNs in the presence of vehicle control dimethy sulfoxide (DMSO) or Bic (25 μM), respectively. (i–vi) Scale bar = 5 μm. (C) The number of GABAergic terminals forming connections with primary dendrites of MSNs (n = 43 dendrites in DMSO/controls, n = 50 dendrites in Bic-treated cultures). (D–F) The number, percentage and size of synaptic α1-, α2- and α1/α2-clusters, respectively, along the primary dendrites of MSNs. The box plots display the median and IQRs of indicated synaptic parameters measured along the first 20 μm of primary dendrites (n = 49 dendrites in DMSO/controls, n = 46 dendrites in Bic-treated cultures) of total of n = 16 neurons from two independent experiments. Statistical analysis was performed using Mann Whitney test, *p < 0.05.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
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Figure 8: GABAA receptor activity has no influence on synapse formation between immature MSNs. (A,B) Immunolabeling of GABAA receptor α1- (i,ii; pink), α2- (iii,iv; green), α1/α2- (v,vi; white) subunit-containing clusters, and presynaptic GABAergic terminals (ii, iv, vi; red) along the primary dendrites of 7 DIV MSNs in the presence of vehicle control dimethy sulfoxide (DMSO) or Bic (25 μM), respectively. (i–vi) Scale bar = 5 μm. (C) The number of GABAergic terminals forming connections with primary dendrites of MSNs (n = 43 dendrites in DMSO/controls, n = 50 dendrites in Bic-treated cultures). (D–F) The number, percentage and size of synaptic α1-, α2- and α1/α2-clusters, respectively, along the primary dendrites of MSNs. The box plots display the median and IQRs of indicated synaptic parameters measured along the first 20 μm of primary dendrites (n = 49 dendrites in DMSO/controls, n = 46 dendrites in Bic-treated cultures) of total of n = 16 neurons from two independent experiments. Statistical analysis was performed using Mann Whitney test, *p < 0.05.
Mentions: To establish whether depolarising activity of GABAA receptors has a regulatory role in synapse formation at the early stage of MSN development, and contrast this with the hyperpolarizing activity of GABAA receptors at the later stage, we have applied a chronic blockade of GABAA receptors using a competitive antagonist Bic (25 μM) in MSN cultures from 4–7 DIV or 7–14 DIV. We have first established that, at this concentration of Bic, the survival of MSNs was not impaired (data not shown). Using the immunocytochemistry approach and analysis described above, we subsequently characterized the formation of GABAergic synapses and the number and size of postsynaptic GABAA receptor clusters (Figures 8A,B).

Bottom Line: When activity of GABAARs was under chronic blockade between 4-7 DIV, the structural properties of these synapses remained unchanged.In contrast, chronic inhibition of GABAARs between 7-14 DIV led to reduction in size of α1- and α1/α2-postsynaptic clusters and a concomitant increase in number and size of α2-postsynaptic clusters.Thus, the main subtypes of GABAergic synapses formed by MSNs are regulated by GABAAR activity, but in opposite directions, and thus appear to be driven by different molecular mechanisms.

View Article: PubMed Central - PubMed

Affiliation: UCL School of Pharmacy, University College London London, UK.

ABSTRACT
Basal ganglia play an essential role in motor coordination and cognitive functions. The GABAergic medium spiny neurons (MSNs) account for ~95% of all the neurons in this brain region. Central to the normal functioning of MSNs is integration of synaptic activity arriving from the glutamatergic corticostriatal and thalamostriatal afferents, with synaptic inhibition mediated by local interneurons and MSN axon collaterals. In this study we have investigated how the specific types of GABAergic synapses between the MSNs develop over time, and how the activity of GABAA receptors (GABAARs) influences this development. Isolated embryonic (E17) MSNs form a homogenous population in vitro and display spontaneous synaptic activity and functional properties similar to their in vivo counterparts. In dual whole-cell recordings of synaptically connected pairs of MSNs, action potential (AP)-activated synaptic events were detected between 7 and 14 days in vitro (DIV), which coincided with the shift in GABAAR operation from depolarization to hyperpolarization, as detected indirectly by intracellular calcium imaging. In parallel, the predominant subtypes of inhibitory synapses, which innervate dendrites of MSNs and contain GABAAR α1 or α2 subunits, underwent distinct changes in the size of postsynaptic clusters, with α1 becoming smaller and α2 larger over time, while both the percentage and the size of mixed α1/α2-postsynaptic clusters were increased. When activity of GABAARs was under chronic blockade between 4-7 DIV, the structural properties of these synapses remained unchanged. In contrast, chronic inhibition of GABAARs between 7-14 DIV led to reduction in size of α1- and α1/α2-postsynaptic clusters and a concomitant increase in number and size of α2-postsynaptic clusters. Thus, the main subtypes of GABAergic synapses formed by MSNs are regulated by GABAAR activity, but in opposite directions, and thus appear to be driven by different molecular mechanisms.

No MeSH data available.


Related in: MedlinePlus