HPLC-Based Activity Profiling for GABA_A Receptor Modulators in Searsia pyroides Using a Larval Zebrafish Locomotor Assay^[*]

Abstract

A dichloromethane extract from leaves of Searsia pyroides potentiated gamma aminobutyric acid-induced chloride currents by 171.8 ± 54% when tested at 100 µg/mL in Xenopus oocytes transiently expressing gamma aminobutyric acid type A receptors composed of α ₁ β ₂ γ ₂s subunits. In zebrafish larvae, the extract significantly lowered pentylenetetrazol-provoked locomotion when tested at 4 µg/mL. Active compounds of the extract were tracked with the aid of HPLC-based activity profiling utilizing a previously validated zebrafish larval locomotor activity assay. From two active HPLC fractions, compounds 1 – 3 were isolated. Structurally related compounds 4 – 6 were purified from a later eluting inactive HPLC fraction. With the aid of ¹H and ¹³C NMR and high-resolution mass spectrometry, compounds 1 – 6 were identified as analogues of anacardic acid. Compounds 1 – 3 led to a concentration-dependent decrease of pentylenetetrazol-provoked locomotion in the zebrafish larvae model, while 4 – 6 were inactive. Compounds 1 – 3 enhanced gamma aminobutyric acid-induced chloride currents in Xenopus oocytes in a concentration-dependent manner, while 4 – 6 only showed marginal enhancements of gamma aminobutyric acid-induced chloride currents. Compounds 2, 3, and 5 have not been reported previously.

Key words

Searsia pyroides - Anacardiaceae - GABA_A receptor - zebrafish larvae - Xenopus oocytes - pentylenetetrazole - HPLC-based activity profiling

• References

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