203 INFLUENCE OF SELF-FERMENTED PINE NEEDLE FRACTIONS ON CATECHOLAMINE RELEASE FROM THE RAT ADRENAL MEDULLA

Abstract

Previously, we have found that self-fermented pine needle (Pinus densiflora) extract inhibits secretion of catecholamines (CA) from the perfused rat adrenal medulla (Shin & Lim, 2010), and also the contractile responses of isolated rat aortic strips induced by phenylephrine and high K+ (Yu et al., 2009). Therefore, the aim of the present study was to investigate the effects of several fractions obtained from methylene chloride (CH2Cl2) extract of self-fermented pine needle on the acetylcholine (ACh)-evoked CA release from the isolated perfused model of the rat adrenal medulla and to establish the mechanism of the most active fraction (Fr.)-induced inhibitory action on the CA release. We obtained 6 fractions from CH2Cl2 extract of self-fermented pine needle. For the ACh (5.32 mM)-evoked CA release, the following rank order of inhibitory potency was obtained: Fr.4>Fr.8-11>>Fr.3>Fr.6=Fr.7>Fr.1.2. Fr.4 (60 μg/mL) perfused into an adrenal vein for 90 min produced relatively time-dependent inhibition of the CA secretory responses to ACh (5.32 mM), high K+ (56 mM), DMPP (100 μM) and McN-A-343 (100 μM). Fr.4 itself did not affect basal CA secretion (data not shown). Also, in the presence of Fr.4 (60 μg/mL), the CA secretory responses to angiotensin II (AngII, 0.1 μM), veratridine (a selective Na+ channel activator, 10 μM), Bay-K-8644 (a L-type dihydropyridine Ca2+ channel activator, 10 μM), and cyclopiazonic acid (a cytoplasmic Ca2+-ATPase inhibitor, 10 μM) were significantly reduced, respectively. In the simultaneous presence of Fr.4 (60 μg/mL) and L-NAME (an inhibitor of NO synthase, 30 μM), the inhibitory responses of Fr.4 on the CA secretion evoked by ACh, high K+, DMPP, McN-A-343, Bay-K-8644, cyclopiazonic acid, AngII and veratridine were considerably recovered to the extent of the corresponding control secretion compared with that of Fr.4-treatment alone. The level of NO released from adrenal medulla after the treatment of Fr.4 (60 μg/mL) was greatly elevated compared with the basal level. Taken together, these results demonstrate that Fr.4 inhibits the CA secretion from the isolated perfused rat adrenal medulla evoked by stimulation of cholinergic receptors as well as by direct membrane-depolarization. It seems that this inhibitory effect of Fr.4 is mediated by blocking the influx of Ca2+ and Na+ into the adrenomedullary chromaffin cells as well as by inhibition of Ca2+ release from the cytoplasmic calcium store, which is evoked at least partly through the increased NO production due to the activation of NO synthase. Based on these results, it is also thought that Fr. 4 isolated from CH2Cl2 extract of pine needle may contain beneficial antihypertensive components to prevent or treat hypertension.