Limits...
Melatonin stimulates dendrite formation and complexity in the hilar zone of the rat hippocampus: participation of the Ca++/Calmodulin complex.

Domínguez-Alonso A, Valdés-Tovar M, Solís-Chagoyán H, Benítez-King G - Int J Mol Sci (2015)

Bottom Line: We found that the CaMKII inhibitor, KN-62, abolished the MEL stimulatory effects on dendritogenesis and that MEL increased the relative amount of CaM in the soluble fraction of hippocampal slices.Our results indicate that MEL stimulates dendrite formation through CaMKII and the translocation of CaM to the soluble fraction.Data strongly suggest that MEL could repair the loss of hippocampal dendrites that occur in neuropsychiatric disorders by increasing CaM levels and activation of CaMKII.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Neurofarmacología, Subdirección de Investigaciones Clínicas, Instituto Nacional de Psiquiatría Ramón de la Fuente Muñiz, Calzada México-Xochimilco No. 101, Col. San Lorenzo-Huipulco, CP 14370 Tlalpan, DF, Mexico. aline.dmgzalonso@gmail.com.

ABSTRACT
Melatonin (MEL), the main product synthesized by the pineal gland, stimulates early and late stages of neurodevelopment in the adult brain. MEL increases dendrite length, thickness and complexity in the hilar and mossy neurons of hippocampus. Dendrite formation involves activation of Ca2+/Calmodulin (CaM)-dependent kinase II (CaMKII) by CaM. Previous work showed that MEL increased the synthesis and translocation of CaM, suggesting that MEL activates CaM-dependent enzymes by this pathway. In this work we investigated whether MEL stimulates dendrite formation by CaMKII activation in organotypic cultures from adult rat hippocampus. We found that the CaMKII inhibitor, KN-62, abolished the MEL stimulatory effects on dendritogenesis and that MEL increased the relative amount of CaM in the soluble fraction of hippocampal slices. Also, PKC inhibition abolished dendritogenesis, while luzindole, an antagonist of MEL receptors (MT1/2), partially blocked the effects of MEL. Moreover, autophosphorylation of CaMKII and PKC was increased in presence of MEL, as well as phosphorylation of ERK1/2. Our results indicate that MEL stimulates dendrite formation through CaMKII and the translocation of CaM to the soluble fraction. Dendritogenesis elicited by MEL also required PKC activation, and signaling through MT1/2 receptors was partially involved. Data strongly suggest that MEL could repair the loss of hippocampal dendrites that occur in neuropsychiatric disorders by increasing CaM levels and activation of CaMKII.

Show MeSH

Related in: MedlinePlus

Effect of Ca2+/CaM-Kinase II on dendrite formation elicited by Melatonin. Participation of CaMKII on dendrite formation elicited by MEL was evaluated by specific inhibition of its activity with KN-62. Thus, rat brain hippocampus was cut in 400 µm slices and cultured in Neurobasal® (GIBCO by Life Technologies, Grand Island, NY, USA) media for 7 days. Then they were incubated for 6 h with either the vehicle (A); 100 nM MEL (B); or pre-incubated with 10 µM KN-62 (C,D) followed by 6 h incubation with the vehicle (C) or 100 nM MEL (D). After the incubation time, slices were cut into 50 µM sections and immunostained for the specific marker of dendrites MAP2. Afterwards, slices were incubated with a secondary antibody coupled to biotin-avidin-peroxidase. Images were acquired with a digital camera coupled to a light microscope with the NIS-Elements software. Scale bar = 100 µm.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4307341&req=5

ijms-16-01907-f001: Effect of Ca2+/CaM-Kinase II on dendrite formation elicited by Melatonin. Participation of CaMKII on dendrite formation elicited by MEL was evaluated by specific inhibition of its activity with KN-62. Thus, rat brain hippocampus was cut in 400 µm slices and cultured in Neurobasal® (GIBCO by Life Technologies, Grand Island, NY, USA) media for 7 days. Then they were incubated for 6 h with either the vehicle (A); 100 nM MEL (B); or pre-incubated with 10 µM KN-62 (C,D) followed by 6 h incubation with the vehicle (C) or 100 nM MEL (D). After the incubation time, slices were cut into 50 µM sections and immunostained for the specific marker of dendrites MAP2. Afterwards, slices were incubated with a secondary antibody coupled to biotin-avidin-peroxidase. Images were acquired with a digital camera coupled to a light microscope with the NIS-Elements software. Scale bar = 100 µm.

Mentions: Previously it was demonstrated that MEL increases dendritogenesis in adult interneurons of the hilar region of the hippocampus and in new neurons formed in the dentate gyrus of rodents [17,18]. Also, it was shown that activation of CaMKII is involved in dendrite formation and maturation [23,44]. Thus, we tested whether the KN-62 compound, a specific inhibitor of CaMKII, suppresses the MEL effects on organotypic hippocampal cultures. As shown in Figure 1A, in presence of the vehicle, neurons of triangular shape with thin dendrites were evident. In presence of 100 nM MEL, neurons developed an intricate web of dendrites (Figure 1B). By contrast, hippocampal slices incubated with the CaMKII inhibitor and the vehicle, showed few and thin dendrites in the interneurons of the hilar zone (Figure 1C), while in hippocampal slices incubated with KN-62 and 100 nM MEL, only triangular and bipolar neurons with short and thin dendrites were evident and the complex dendrite network elicited by the indoleamine was not observed (Figure 1D). Since MEL-induced dendrite formation was abolished in the presence of the CaMKII inhibitor, data indicate that the enzyme participates in the mechanism by which MEL elicited dendritogenesis.


Melatonin stimulates dendrite formation and complexity in the hilar zone of the rat hippocampus: participation of the Ca++/Calmodulin complex.

Domínguez-Alonso A, Valdés-Tovar M, Solís-Chagoyán H, Benítez-King G - Int J Mol Sci (2015)

Effect of Ca2+/CaM-Kinase II on dendrite formation elicited by Melatonin. Participation of CaMKII on dendrite formation elicited by MEL was evaluated by specific inhibition of its activity with KN-62. Thus, rat brain hippocampus was cut in 400 µm slices and cultured in Neurobasal® (GIBCO by Life Technologies, Grand Island, NY, USA) media for 7 days. Then they were incubated for 6 h with either the vehicle (A); 100 nM MEL (B); or pre-incubated with 10 µM KN-62 (C,D) followed by 6 h incubation with the vehicle (C) or 100 nM MEL (D). After the incubation time, slices were cut into 50 µM sections and immunostained for the specific marker of dendrites MAP2. Afterwards, slices were incubated with a secondary antibody coupled to biotin-avidin-peroxidase. Images were acquired with a digital camera coupled to a light microscope with the NIS-Elements software. Scale bar = 100 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4307341&req=5

ijms-16-01907-f001: Effect of Ca2+/CaM-Kinase II on dendrite formation elicited by Melatonin. Participation of CaMKII on dendrite formation elicited by MEL was evaluated by specific inhibition of its activity with KN-62. Thus, rat brain hippocampus was cut in 400 µm slices and cultured in Neurobasal® (GIBCO by Life Technologies, Grand Island, NY, USA) media for 7 days. Then they were incubated for 6 h with either the vehicle (A); 100 nM MEL (B); or pre-incubated with 10 µM KN-62 (C,D) followed by 6 h incubation with the vehicle (C) or 100 nM MEL (D). After the incubation time, slices were cut into 50 µM sections and immunostained for the specific marker of dendrites MAP2. Afterwards, slices were incubated with a secondary antibody coupled to biotin-avidin-peroxidase. Images were acquired with a digital camera coupled to a light microscope with the NIS-Elements software. Scale bar = 100 µm.
Mentions: Previously it was demonstrated that MEL increases dendritogenesis in adult interneurons of the hilar region of the hippocampus and in new neurons formed in the dentate gyrus of rodents [17,18]. Also, it was shown that activation of CaMKII is involved in dendrite formation and maturation [23,44]. Thus, we tested whether the KN-62 compound, a specific inhibitor of CaMKII, suppresses the MEL effects on organotypic hippocampal cultures. As shown in Figure 1A, in presence of the vehicle, neurons of triangular shape with thin dendrites were evident. In presence of 100 nM MEL, neurons developed an intricate web of dendrites (Figure 1B). By contrast, hippocampal slices incubated with the CaMKII inhibitor and the vehicle, showed few and thin dendrites in the interneurons of the hilar zone (Figure 1C), while in hippocampal slices incubated with KN-62 and 100 nM MEL, only triangular and bipolar neurons with short and thin dendrites were evident and the complex dendrite network elicited by the indoleamine was not observed (Figure 1D). Since MEL-induced dendrite formation was abolished in the presence of the CaMKII inhibitor, data indicate that the enzyme participates in the mechanism by which MEL elicited dendritogenesis.

Bottom Line: We found that the CaMKII inhibitor, KN-62, abolished the MEL stimulatory effects on dendritogenesis and that MEL increased the relative amount of CaM in the soluble fraction of hippocampal slices.Our results indicate that MEL stimulates dendrite formation through CaMKII and the translocation of CaM to the soluble fraction.Data strongly suggest that MEL could repair the loss of hippocampal dendrites that occur in neuropsychiatric disorders by increasing CaM levels and activation of CaMKII.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Neurofarmacología, Subdirección de Investigaciones Clínicas, Instituto Nacional de Psiquiatría Ramón de la Fuente Muñiz, Calzada México-Xochimilco No. 101, Col. San Lorenzo-Huipulco, CP 14370 Tlalpan, DF, Mexico. aline.dmgzalonso@gmail.com.

ABSTRACT
Melatonin (MEL), the main product synthesized by the pineal gland, stimulates early and late stages of neurodevelopment in the adult brain. MEL increases dendrite length, thickness and complexity in the hilar and mossy neurons of hippocampus. Dendrite formation involves activation of Ca2+/Calmodulin (CaM)-dependent kinase II (CaMKII) by CaM. Previous work showed that MEL increased the synthesis and translocation of CaM, suggesting that MEL activates CaM-dependent enzymes by this pathway. In this work we investigated whether MEL stimulates dendrite formation by CaMKII activation in organotypic cultures from adult rat hippocampus. We found that the CaMKII inhibitor, KN-62, abolished the MEL stimulatory effects on dendritogenesis and that MEL increased the relative amount of CaM in the soluble fraction of hippocampal slices. Also, PKC inhibition abolished dendritogenesis, while luzindole, an antagonist of MEL receptors (MT1/2), partially blocked the effects of MEL. Moreover, autophosphorylation of CaMKII and PKC was increased in presence of MEL, as well as phosphorylation of ERK1/2. Our results indicate that MEL stimulates dendrite formation through CaMKII and the translocation of CaM to the soluble fraction. Dendritogenesis elicited by MEL also required PKC activation, and signaling through MT1/2 receptors was partially involved. Data strongly suggest that MEL could repair the loss of hippocampal dendrites that occur in neuropsychiatric disorders by increasing CaM levels and activation of CaMKII.

Show MeSH
Related in: MedlinePlus