8-bromocyclic-gmp and osthol

Osthole, an active constituent isolated from Cnidium monnieri (L.) Cusson, has previously been shown to have the capacity to increase depolarization-evoked glutamate release in rat hippocampal nerve terminals. As cGMP-dependent signaling cascade has been found to modulate glutamate release at the presynaptic level, the aim of this study was to further examine the role of cGMP signaling pathway in the regulation of osthole on glutamate release in hippocampal synaptosomes. Results showed that osthole dose-dependently increased intrasynaptosomal cGMP levels. The elevation of cGMP levels by osthole was prevented by the phosphodiesterase 5 inhibitor sildenafil but was insensitive to the guanylyl cyclase inhibitor ODQ. In addition, osthole-induced facilitation of 4-aminopyridine (4-AP)-evoked glutamate release was completely prevented by the cGMP-dependent protein kinase (PKG) inhibitors, KT5823, and Rp-8-Br-PET-cGMPS. Direct activation of PKG with 8-Br-cGMP or 8-pCPT-cGMP also occluded the osthole-mediated facilitation of 4-AP-evoked glutamate release. Furthermore, sildenafil exhibited a dose-dependent facilitation of 4-AP-evoked release of glutamate and occluded the effect of osthole on the 4-AP-evoked glutamate release. Collectively, our findings suggest that osthole-mediated facilitation of glutamate release involves the activation of cGMP/PKG-dependent pathway.

Topics: Animals; Carbazoles; Coumarins; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Cyclic Nucleotide Phosphodiesterases, Type 5; Glutamic Acid; Guanylate Cyclase; Hippocampus; Male; Nerve Endings; Rats; Rats, Sprague-Dawley; Synaptic Transmission; Thionucleotides

The effects of osthole, a coumarin isolated from Cnidium monnieri (L.) Cusson, on ionic currents in a mouse neuroblastoma and rat glioma hybrid cell line, NG105-18, were investigated with the aid of the whole-cell voltage-clamp technique. Osthole (0.3-100 microM) caused an inhibition of voltage-dependent L-type Ca(2+) current (I(Ca,L)) in a concentration-dependent manner. Osthole produced no change in the overall shape of the current-voltage relationship of I(Ca,L). The IC(50) value of the osthole-induced inhibition of I(Ca,L) was 4 microM. The presence of osthole (3 microM) shifted the steady state inactivation curve of I(Ca,L) to a more negative potential by approximately -15mV. Osthole (3 microM) also produced a prolongation in the recovery of I(Ca,L) inactivation. Although osthole might suppress phosophodiesterases to increase intracellular adenosine-3',5'-cyclic monophosphate (cyclic AMP) or guanosine-3',5'-cyclic monophosphate (cyclic GMP), sp-cAMPS did not affect I(Ca,L) and 8-bromo-cyclic GMP slightly suppressed it. Thus, osthole-mediated inhibition of I(Ca,L) was not associated with intracellular cyclic AMP or GMP. However, no effect of osthole on voltage-dependent K(+) outward current was observed. Under a current-clamp mode, osthole could decrease the firing frequency of action potentials. Therefore, the channel-blocking properties of osthole may, at least in part, contribute to the underlying mechanisms by which it affects neuronal or neuroendocrine function.

Topics: Animals; Calcium Channel Blockers; Calcium Channels, L-Type; Coumarins; Cyclic AMP; Cyclic GMP; Hybrid Cells; Membrane Potentials; Mice; Neurons; Nimodipine; omega-Conotoxin GVIA; Potassium Channels, Voltage-Gated; Rats; Thionucleotides