Rapid actions of estradiol on cyclic amp response-element binding protein phosphorylation in dorsal root ganglion neurons
Section snippets
Neuronal cultures
Adult outbred Wistar rats (200–300 g) were used for the experiments. All experiments were performed in accordance with the National Health and Medical Research Council of Australia's Code of Practice for the Care and Use of Animals for Experimental Purposes, which also conforms to standard international guidelines. Every attempt was made to reduce the number of animals used and to minimise their suffering. Animals were deeply anesthetized (sodium pentobarbitone, 48 mg/kg, i.p.) prior to
Estradiol activates ERK in primary sensory neurons
Western blot analysis of cultured DRG neurons showed that treatment with estradiol (10 nM) induced phosphorylation of p44 (ERK1) and p42 ERK (ERK2) by 10 min and this was still elevated at 60 min (Fig. 1A). Data have been normalized to 10 min control data. The 30 and 60 min controls decrease relative to the 10 min control because ERK activation occurring in response to the action of treating the cells (movement of culture dish, temperature change, light) decreases rapidly over this period (Fig.
Discussion
CREB is a critical molecule associated with neuronal plasticity and adaptation, and CREB activation via ERK and PI3-K is a key target of neurotrophin-activated signaling pathways (Shaywitz and Greenberg, 1999). We visualized CREB activation in cultures of adult DRG and identified individual sensory neurons that respond rapidly to estradiol in vitro. In these neurons, estradiol exposure resulted in CREB phosphorylation and this occurred via activation of the ERK pathway and did not involve the
Acknowledgments
This work was supported by the National Health and Medical Research Council (Australia), Project grant 990034 and Senior Research Fellowship 157213, and University of New South Wales Faculty Research grant to J.R.K. We are grateful to Peregrine Osborne for valuable discussions.
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