Identification and characterization of GABA(A) receptor modulatory diterpenes from Biota orientalis that decrease locomotor activity in mice.

An ethyl acetate extract of Biota orientalis leaves potentiated GABA-induced control current by 92.6% +/- 22.5% when tested at 100 mu g/mL in Xenopus laevis oocytes expressing GABA(A) receptors (alpha(1)beta(2)gamma(2S) subtype) in two-microelectrode voltage clamp measurements. HPLC-based activity profiling was used to identify isopimaric acid (4) and sandaracopimaric acid (5) as the compounds largely responsible for the activity. Sandaracopimaradienolal (3) was characterized as a new natural product. Compounds 4 and 5 were investigated for GABAA receptor subtype selectivity at the subtypes alpha(1)beta(1)gamma(2S), alpha(1)beta(2)gamma(2S), alpha(1)beta(3)gamma(2S), alpha(2)beta(2)gamma(2S), alpha(3)beta(2)gamma(2S), and alpha(5)beta(2)gamma(2S). Sandaracopimaric acid (5) was significantly more potent than isopimaric acid (4) at the GABA(A) receptor subtypes alpha(1)beta(1)gamma(2S), alpha(2)beta(2)gamma(2S), and alpha(5)beta(2)gamma(2S) (EC(50) 4: 289.5 +/- 82.0, 364.8 +/- 85.0, and 317.0 +/- 83.7 mu M vs EC(50) 5: 48.1 +/- 13.4, 31.2 +/- 4.8, and 40.7 +/- 14.7 mu M). The highest efficiency was reached by 4 and 5 on alpha(2)- and alpha(3)-containing receptor subtypes. In the open field test, ip administration of 5 induced a dose-dependent decrease of locomotor activity in a range of 3 to 30 mg/kg body weight in mice. No significant anxiolytic-like activity was observed in doses between 1 and 30 mg/kg body weight in mice.