1. ¹Key Laboratory of Reproductive Medicine, Department of Pharmacology, Nanjing Medical University, Nanjing, PR China
2. ²Department of Geriatrics, the First Affiliated Hospital of Nanjing Medical University, Nanjing, PR China
3. ³Ion Channel & Transporter Laboratory, Department of Physiology, National University of Singapore, Singapore

Correspondence: Dr S Li, Department of Pharmacology, Nanjing Medical University, Hanzhong Road 140, Nanjing 210029, PR China. Tel: +86 25 86863364; Fax: +86 25 86863050; E-mail: snli@njmu.edu.cn

Received 14 December 2005; Revised 18 April 2006; Accepted 4 May 2006; Published online 7 June 2006.

Abstract

Urocortin 2, a new member of the corticotrophin-releasing factor (CRF) neuropeptide family, was reported to be widely expressed in the central nervous system and peripheral tissues. Here, we detected urocortin 2 mRNA in PC12 cells using reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, we observed its effects on intracellular Ca²⁺ concentration ([Ca²⁺]_i) using confocal microscopy and flow cytometry and on voltage-gated calcium channel (VGCC) currents using whole-cell patch clamp. Our results showed that urocortin 2 mRNA was coexpressed with CRF, and CRF receptor (CRFR) 2 in undifferentiated PC12 cells, but not CRFR1 or CRFR2. KCl (40 mM) or Bay K8644 (1 M), an L-type VGCC activator, increased [Ca²⁺]_i. Pretreatment of the cells with urocortin 2 significantly diminished the effect of Bay K8644 or KCl. Urocortin 2 showed no influence on [Ca²⁺]_i in tyrode's solution containing EGTA or Ca²⁺-free tyrode's solution. It reversibly inhibited the VGCC currents in a concentration-dependent manner, but had no apparent effects on the cells treated with nifedipine (1 M), an L-type VGCC blocker. Urocortin 2 up-shifted the current–voltage curves. No frequency-dependence of urocortin 2 effects on I_Ba was observed. The inhibitory effects of urocortin 2 on VGCC currents or [Ca²⁺]_i were not affected by astressin 2B, an antagonist of CRFR2. As calcium overload play a key role in some neuronal degenerative diseases such as Alzheimer's and Parkinson's diseases, our results suggest that urocortin 2 may be a potentially interesting agent for the treatment of these diseases.