WILLARDIINES DIFFERENTIATE AGONIST BINDING-SITES FOR KAINATE-PREFERRING VERSUS AMPA-PREFERRING GLUTAMATE RECEPTORS IN DRG-NEURONS AND HIPPOCAMPAL-NEURONS

Concentration jump responses to ti-substituted (S)-willardiines were r ecorded from dorsal root ganglion (DRG) and hippocampal neurons under voltage clamp. After block of desensitization by concanavalin-A, dose- response analysis for activation of kainate-preferring receptors in DR G neurons gave the potency sequence trifluoromethyl > iodo > bromo app roximate to chloro > nitro approximate to cyano > kainate > methyl > f luoro > (R,S)-AMPA >> willardiine; EC(50) values for the most and leas t potent willardiine derivatives, 5-trifluoromethyl (70 nM) and 5-fluo ro (69 mu M), differed 1000-fold. The potency sequence for equilibrium responses at AMPA-preferring receptors in hippocampal neurons was str ikingly different from that obtained in DRG neurons: fluoro > cyano ap proximate to trifluoromethyl approximate to nitro > chloro approximate to bromo > (R,S)-AMPA > iodo > willardiine > kainate > methyl. In hip pocampal neurons EC(50) values for the most and least potent willardii ne derivatives, 5-fluoro (1.5 mu M) and 5-methyl (251 mu M), differed only 170-fold. Consistent with equilibrium potency measurements, in DR G neurons the kinetics of deactivation for willardiines, recorded foll owing a return to agonist-free solution, were rapid for 5-fluoro (tau( off) = 43 msec) but slow for 5-iodo (tau(off) = 4.2 sec), while the op posite sequence was observed for hippocampal neurons, slow for 5-fluor o (tau(off) = 2.1 sec) and rapid for 5-iodo (tau(off) = 188 msec). The kinetics of recovery from desensitization showed comparable agonist- and cell-dependent differences. Structure-activity analysis for agonis t responses recorded from DRG and hippocampal neurons suggests that fo r both kainate-preferring and AMPA-preferring receptors the binding of willardiines involves interactions with polar groups such that potenc y is related to ionization of the uracil ring, and hence the electron- withdrawing ability of the 5-position substituent. However, kainate-pr eferring receptors differ from AMPA-preferring receptors in possessing a lipophilic pocket that further enhances agonist potency by hydropho bic bonding of the 5-substituent. In contrast, AMPA-preferring recepto rs lack such a lipophilic site, and for 5-position substituents of the same electron-withdrawing ability, potency decreases with increase in size.