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Role for MMP-9 in stress-induced downregulation of nectin-3 in hippocampal CA1 and associated behavioural alterations.

van der Kooij MA, Fantin M, Rejmak E, Grosse J, Zanoletti O, Fournier C, Ganguly K, Kalita K, Kaczmarek L, Sandi C - Nat Commun (2014)

Bottom Line: A reduction in nectin-3 in the perisynaptic CA1, but not in the CA3, compartment is observed following chronic stress and is implicated in the effects of stress in social exploration, social recognition and a CA1-dependent cognitive task.Increased MMP-9-related gelatinase activity, involving N-methyl-D-aspartate receptor, is specifically found in the CA1 and involved in nectin-3 cleavage and chronic stress-induced social and cognitive alterations.Thus, MMP-9 proteolytic processing emerges as an important mediator of stress effects in brain function and behaviour.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Behavioral Genetics, Brain Mind Institute, School of Life Sciences, École Polytechnique Fédérale de Lausanne, EPFL, Lausanne 1015, Switzerland.

ABSTRACT
Chronic stress is a risk factor for the development of psychopathologies characterized by cognitive dysfunction and deregulated social behaviours. Emerging evidence suggests a role for cell adhesion molecules, including nectin-3, in the mechanisms that underlie the behavioural effects of stress. We tested the hypothesis that proteolytic processing of nectins by matrix metalloproteinases (MMPs), an enzyme family that degrades numerous substrates, including cell adhesion molecules, is involved in hippocampal effects induced by chronic restraint stress. A reduction in nectin-3 in the perisynaptic CA1, but not in the CA3, compartment is observed following chronic stress and is implicated in the effects of stress in social exploration, social recognition and a CA1-dependent cognitive task. Increased MMP-9-related gelatinase activity, involving N-methyl-D-aspartate receptor, is specifically found in the CA1 and involved in nectin-3 cleavage and chronic stress-induced social and cognitive alterations. Thus, MMP-9 proteolytic processing emerges as an important mediator of stress effects in brain function and behaviour.

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Nectin-3 cleavage is induced by glutamate and is NMDA- and MMP-9 dependent.Data in a–c are derived from three independent experiments. (a,b) Hippocampal neurons at day 7 were stimulated in vitro with KCl, glutamate or NMDA for 30 min in the presence or absence of inhibitors. Cleavage of full nectin-3 (83 kDa) results in the accumulation of a SPF (small proteolytic fragment). Nectin-3 cleavage was observed after glutamate and NMDA, whereas NMDA-induced nectin-3 shedding was inhibited in the presence of the NMDA-inhibitor APV (a: F2,6=6.25, P=0.04; b: F2,14=15.05, P<0.001). (c) Incubation of the hippocampal neurons with a MMP-9 inhibitor abolished the NMDA-induced nectin-3 cleavage (F2,13=13.76, P<0.001). (d) In synaptoneurosomes obtained from the CA1 from controls and stressed animals, we found that glutamate-induced nectin-3 shedding was more pronounced in stressed animals and that this effect could be prevented by application of a specific MMP-9 inhibitor (effect of stress: F1,28=33.71, P<0.0001; effect of MMP-9 inhibition: F1,28=7.797, P=0.0093, n=8 per group). (e) Cleavage of recombinant nectin-3 by MMP-9 in vitro. Recombinant nectin-3 was incubated with purified recombinant non-active MMP-9 (E402A), autoactivating MMP-9, autoactivating MMP-9 in the presence of 5 μM MMP-9 inhibitor, or no proteinase (buffer) as indicated. The digestion products were resolved by western blot analysis and probed with anti-His-Tag antibody. Error bars represent s.e.m. *P<0.05, **P<0.01 (one- or two-way analysis of variance followed by Bonferroni post hoc comparisons, where applicable).
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f4: Nectin-3 cleavage is induced by glutamate and is NMDA- and MMP-9 dependent.Data in a–c are derived from three independent experiments. (a,b) Hippocampal neurons at day 7 were stimulated in vitro with KCl, glutamate or NMDA for 30 min in the presence or absence of inhibitors. Cleavage of full nectin-3 (83 kDa) results in the accumulation of a SPF (small proteolytic fragment). Nectin-3 cleavage was observed after glutamate and NMDA, whereas NMDA-induced nectin-3 shedding was inhibited in the presence of the NMDA-inhibitor APV (a: F2,6=6.25, P=0.04; b: F2,14=15.05, P<0.001). (c) Incubation of the hippocampal neurons with a MMP-9 inhibitor abolished the NMDA-induced nectin-3 cleavage (F2,13=13.76, P<0.001). (d) In synaptoneurosomes obtained from the CA1 from controls and stressed animals, we found that glutamate-induced nectin-3 shedding was more pronounced in stressed animals and that this effect could be prevented by application of a specific MMP-9 inhibitor (effect of stress: F1,28=33.71, P<0.0001; effect of MMP-9 inhibition: F1,28=7.797, P=0.0093, n=8 per group). (e) Cleavage of recombinant nectin-3 by MMP-9 in vitro. Recombinant nectin-3 was incubated with purified recombinant non-active MMP-9 (E402A), autoactivating MMP-9, autoactivating MMP-9 in the presence of 5 μM MMP-9 inhibitor, or no proteinase (buffer) as indicated. The digestion products were resolved by western blot analysis and probed with anti-His-Tag antibody. Error bars represent s.e.m. *P<0.05, **P<0.01 (one- or two-way analysis of variance followed by Bonferroni post hoc comparisons, where applicable).

Mentions: Stress increases extracellular glutamate levels in the hippocampus41. To determine whether nectin-3 cleavage may result from enhanced neuronal activity, cultured hippocampal neurons were treated with either 30 mM KCl or 50 μM glutamate and extracts from whole-cell lysates were analysed via western immunoblotting. We found that glutamate treatment resulted in a significant increase in the level of the cleaved form of nectin-3 (small proteolytic fragment; Fig. 4a). This cleavage was dependent on NMDA receptor function, as it was reproduced by treatment with 100 μM NMDA and could be entirely inhibited by the NMDA receptor antagonists APV (DL-2-amino-5-phosphonopentaoic acid) or MK-801 (Fig. 4b).


Role for MMP-9 in stress-induced downregulation of nectin-3 in hippocampal CA1 and associated behavioural alterations.

van der Kooij MA, Fantin M, Rejmak E, Grosse J, Zanoletti O, Fournier C, Ganguly K, Kalita K, Kaczmarek L, Sandi C - Nat Commun (2014)

Nectin-3 cleavage is induced by glutamate and is NMDA- and MMP-9 dependent.Data in a–c are derived from three independent experiments. (a,b) Hippocampal neurons at day 7 were stimulated in vitro with KCl, glutamate or NMDA for 30 min in the presence or absence of inhibitors. Cleavage of full nectin-3 (83 kDa) results in the accumulation of a SPF (small proteolytic fragment). Nectin-3 cleavage was observed after glutamate and NMDA, whereas NMDA-induced nectin-3 shedding was inhibited in the presence of the NMDA-inhibitor APV (a: F2,6=6.25, P=0.04; b: F2,14=15.05, P<0.001). (c) Incubation of the hippocampal neurons with a MMP-9 inhibitor abolished the NMDA-induced nectin-3 cleavage (F2,13=13.76, P<0.001). (d) In synaptoneurosomes obtained from the CA1 from controls and stressed animals, we found that glutamate-induced nectin-3 shedding was more pronounced in stressed animals and that this effect could be prevented by application of a specific MMP-9 inhibitor (effect of stress: F1,28=33.71, P<0.0001; effect of MMP-9 inhibition: F1,28=7.797, P=0.0093, n=8 per group). (e) Cleavage of recombinant nectin-3 by MMP-9 in vitro. Recombinant nectin-3 was incubated with purified recombinant non-active MMP-9 (E402A), autoactivating MMP-9, autoactivating MMP-9 in the presence of 5 μM MMP-9 inhibitor, or no proteinase (buffer) as indicated. The digestion products were resolved by western blot analysis and probed with anti-His-Tag antibody. Error bars represent s.e.m. *P<0.05, **P<0.01 (one- or two-way analysis of variance followed by Bonferroni post hoc comparisons, where applicable).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4199199&req=5

f4: Nectin-3 cleavage is induced by glutamate and is NMDA- and MMP-9 dependent.Data in a–c are derived from three independent experiments. (a,b) Hippocampal neurons at day 7 were stimulated in vitro with KCl, glutamate or NMDA for 30 min in the presence or absence of inhibitors. Cleavage of full nectin-3 (83 kDa) results in the accumulation of a SPF (small proteolytic fragment). Nectin-3 cleavage was observed after glutamate and NMDA, whereas NMDA-induced nectin-3 shedding was inhibited in the presence of the NMDA-inhibitor APV (a: F2,6=6.25, P=0.04; b: F2,14=15.05, P<0.001). (c) Incubation of the hippocampal neurons with a MMP-9 inhibitor abolished the NMDA-induced nectin-3 cleavage (F2,13=13.76, P<0.001). (d) In synaptoneurosomes obtained from the CA1 from controls and stressed animals, we found that glutamate-induced nectin-3 shedding was more pronounced in stressed animals and that this effect could be prevented by application of a specific MMP-9 inhibitor (effect of stress: F1,28=33.71, P<0.0001; effect of MMP-9 inhibition: F1,28=7.797, P=0.0093, n=8 per group). (e) Cleavage of recombinant nectin-3 by MMP-9 in vitro. Recombinant nectin-3 was incubated with purified recombinant non-active MMP-9 (E402A), autoactivating MMP-9, autoactivating MMP-9 in the presence of 5 μM MMP-9 inhibitor, or no proteinase (buffer) as indicated. The digestion products were resolved by western blot analysis and probed with anti-His-Tag antibody. Error bars represent s.e.m. *P<0.05, **P<0.01 (one- or two-way analysis of variance followed by Bonferroni post hoc comparisons, where applicable).
Mentions: Stress increases extracellular glutamate levels in the hippocampus41. To determine whether nectin-3 cleavage may result from enhanced neuronal activity, cultured hippocampal neurons were treated with either 30 mM KCl or 50 μM glutamate and extracts from whole-cell lysates were analysed via western immunoblotting. We found that glutamate treatment resulted in a significant increase in the level of the cleaved form of nectin-3 (small proteolytic fragment; Fig. 4a). This cleavage was dependent on NMDA receptor function, as it was reproduced by treatment with 100 μM NMDA and could be entirely inhibited by the NMDA receptor antagonists APV (DL-2-amino-5-phosphonopentaoic acid) or MK-801 (Fig. 4b).

Bottom Line: A reduction in nectin-3 in the perisynaptic CA1, but not in the CA3, compartment is observed following chronic stress and is implicated in the effects of stress in social exploration, social recognition and a CA1-dependent cognitive task.Increased MMP-9-related gelatinase activity, involving N-methyl-D-aspartate receptor, is specifically found in the CA1 and involved in nectin-3 cleavage and chronic stress-induced social and cognitive alterations.Thus, MMP-9 proteolytic processing emerges as an important mediator of stress effects in brain function and behaviour.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Behavioral Genetics, Brain Mind Institute, School of Life Sciences, École Polytechnique Fédérale de Lausanne, EPFL, Lausanne 1015, Switzerland.

ABSTRACT
Chronic stress is a risk factor for the development of psychopathologies characterized by cognitive dysfunction and deregulated social behaviours. Emerging evidence suggests a role for cell adhesion molecules, including nectin-3, in the mechanisms that underlie the behavioural effects of stress. We tested the hypothesis that proteolytic processing of nectins by matrix metalloproteinases (MMPs), an enzyme family that degrades numerous substrates, including cell adhesion molecules, is involved in hippocampal effects induced by chronic restraint stress. A reduction in nectin-3 in the perisynaptic CA1, but not in the CA3, compartment is observed following chronic stress and is implicated in the effects of stress in social exploration, social recognition and a CA1-dependent cognitive task. Increased MMP-9-related gelatinase activity, involving N-methyl-D-aspartate receptor, is specifically found in the CA1 and involved in nectin-3 cleavage and chronic stress-induced social and cognitive alterations. Thus, MMP-9 proteolytic processing emerges as an important mediator of stress effects in brain function and behaviour.

Show MeSH
Related in: MedlinePlus