MDMA fails to potentiate kainate-induced neurotoxicity but increases sensitivity to subconvulsant doses of this glutamatergic agonist

S. Abad1, D. Pubill1, M. Pallas1, C. Auladell2, J. Camarasa1, A. Camins1, F. Junyent1, E. Escubedo1. 1University of Barcelona, Pharmacology and Therapeutic Chemistry, 08028, Spain, 2University of Barcelona, Cellular Biology, 08028, Spain

In previous studies, our group showed that MDMA produces a sustained increase of cytosolic calcium levels along 24 hours that was mediated by a partial agonist effect on α-7 nicotinic receptors. Based on these results we hypothesized that MDMA administered in vivo may modify the levels of intracellular calcium, exacerbating the kainate-induced neurotoxicity which is calcium mediated.

Male C57BL/6 mice (n=58 )one month old were treated using the following schedule:

MDMA: 3 s.c. doses/day every 3 h, once per week for 4 weeks. Doses increased weekly (5 mg/kg – 7.5mg/kg – 7.5mg/kg – 10mg/kg); K20: Kainate 20 mg/kg i.p a single dose; K30: Kainate 30 mg/kg i.p a single dose; MK20 and MK30: MDMA treatment was the same that MDMA group and kainate (20 mg/kg or 30 mg/kg) was administered 24h post last MDMA dose. One-way analysis of variance (ANOVA) was used to determine overall treatment effects on seizures. During the following 3h post kainate administration, seizure severity was scored on a six-point behavioral own scale. 24 o 72h later all animals were sacrificed and perfused with 4% paraformaldehyde. After equilibration with 30% sucrose solution, brains were frozen and stored. Detection of calbindin, GFAP and microglia were carried out by immunohistochemistry. To reveal degenerative neurons, FluoroJade B staining was applied. Animals of MDMA group did not show any seizure. As expected, all K30 animals showed seizures. The seizure index was greater than that of K20. MDMA potentiated K30 and K20-induced seizures. Moreover, the latency period was shorter and lasted for a longer period in MDMA pre-treated mice.In MK30 group, 25% of animals died throughout 24h post kainate administration.

The effect on the cytosolic calcium levels identified in vitro has been reflected in vivo by changes in calbindin expression, a calcium buffer protein. Similarly to kainate, MDMA showed a biphasic effect on the calbindin expression, producing a decrease in the labeling of hippocampal CA1 neurons and in the dentate gyrus granular neurons, which back to basal levels 48h later. It must be pointed that final calbindin levels in MDMA group were greater than controls. In addition, MK30 group levels followed the same profile than MDMA itself. No dead or dying cells in any brain region of groups MDMA, K20 and MK20 were detected using the FluoroJade B. Group of animals injected with kainate 30 mg/kg showed a positive labeling in hippocampal CA1, CA3 and in the dentate gyrus, which was not increased by previous MDMA treatment. Labeling astrogliosis with GFAP revealed a significant increase in group K30 that was potentiated by previous MDMA treatment. In conclusion, MDMA potentiates the kainate lethality, decreases the cell threshold of depolarization down to the point that animals become more sensitive to a subconvulsant dose of kainate, but did not potentiate kainate-mediated neurotoxicity although more activated astroglia is revealed.

This work was supported by grants from Ministerio de Ciencia e Innovación (SAF2010-15948); Plan Nacional sobre Drogas (2010/005) and Generalitat de Catalunya (SGR977 and 2009/SGR977 and SGR00853). Also by grants BFU2010-19119/BFI, and grant 610RT0405 from CYTED.