Biochemical and Biophysical Research Communications
TRPV1 stimulation triggers apoptotic cell death of rat cortical neurons
Section snippets
Materials and methods
Preparation of neuron-rich cultures from rat cerebral cortex. All animals used in the present study were treated in accordance with the guidelines of the Kyoto University Animal Experimentation Committee and the Japanese Pharmacological Society. Primary cultures of dissociated cortical neurons were prepared from the cerebral cortex of fetal Wistar rats (18 days of gestation), according to procedures described previously [13], [14]. Briefly, single cells mechanically dissociated from the
Functional expression of TRPV1 in rat cortical cultures
We examined whether cultured rat cortical cells expressed functional TRPV1. RT-PCR and Western blot analyses revealed that TRPV1 is expressed at the mRNA and protein level after 11-13 days in culture (Fig. 1A and B). Furthermore, calcium imaging with fura-2 AM demonstrated that a TRPV1-specific agonist, capsaicin (10 μM), can induce an increase in ([Ca2+]i) that was markedly attenuated by a TRPV1-specific antagonist, capsazepine (10 μM). This suggests that TRPV1 is activated in response to
Discussion
Here, we found that overstimulation of TRPV1 in rat cortical neurons led to apoptotic cell death via ERK phosphorylation, L-type VDCC opening, ROS production and caspase-3 activation, which agrees with previous studies showing that capsaicin can induce apoptotic cell death in rat mesencephalic dopamine neurons [11], [12].
We found that NADA, which is an endogenous TRPV1 agonist, concentration-dependently exert a neurotoxic effect on cortical neurons. An antagonist of TRPV1 largely inhibited
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2018, Life SciencesCitation Excerpt :Our data also show that AM404 induces a TRPV1-dependent cytotoxicity both in HEK 293 cells expressing hTRPV1 and in rat DRG neurons. Activation of TRPV1 by capsaicin and other agonists mediates cytotoxicity in several rodent and human cellular systems, including human neuronal tissue [16,19–23,32–37]. For assessment of cytotoxic properties of AM404 in the stable hTRPV1-HEK 293 cells we used a commonly applied flow cytometric assay for detection of cell death.