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The effects of extracellularly-applied synthetic philanthotoxin-343 (PhTX-343) on transmission and long-term potentiation (LTP) at Schaffer-collateral/commissural-CA1 synapses were investigated. PhTX-343 was ineffective in antagonizing CA1 field-EPSPs mediated by AMPA/kainate receptors. However, when a micromolar concentration of the toxin was present during tetanization, the induction of LTP was suppressed. In contrast, when PhTX-343 was applied either immediately after or long after tetanization no effect on LTP could be found. It appears that the synaptic, non-NMDA receptors of the CA1-region are insensitive to PhTX-343. Suppression of LTP induction could result from antagonism of postsynaptic NMDA receptors, but the results do not rule out other possibilities such as presynaptic block.

We have purified and characterized two vertebrate excitatory amino acid ionotropic receptors from the Xenopus central nervous system. Each is a unitary receptor (i.e., having more than one class of excitatory amino acid agonist specificity within one protein oligomer). The first is a unitary non-N-methyl-D-aspartate (non-NMDA) receptor and the second is a unitary NMDA/non-NMDA receptor. The specific agonist-activated channel activity and pharmacology of each type were recognized by patch-clamping lipid bilayers in which the isolated protein was reconstituted. In the second case, the NMDA and the non-NMDA sites could not be physically separated and exhibited functional interaction. Parallel evidence for this was obtained when poly(A) RNA from Xenopus brain was translated in oocytes: a noncompetitive inhibition of the response to L-kainate is produced by NMDA to a maximum depression of 30% at 1 mM NMDA. Each isolated oligomer contains 42-kDa subunits of the non-NMDA ligand binding type, but the second type has an additional NMDA-receptor-specific 100-kDa subunit. Thus, a subunit-exchange hypothesis can account for the known multiplicity of excitatory amino acid receptor types.

The effects of spermine and a synthetic analogue (PhTX-343) of the polyamine amide toxin, delta-philanthotoxin, on the responses of Xenopus oocytes to application of amino acids were examined using voltage clamp. The oocytes were injected with either total rat brain RNA or chick cerebrum, poly(A+)RNA. The responses to N-methyl-D-aspartate and L-kainate were potentiated by low concentrations (10(-11)-10(-7) M) of PhTX-343 and by 10(-5)-10(-4) M spermine. There was variability between oocytes in terms of their responsiveness to these compounds and recovery from their effects was slow and often incomplete. Prolonged or repeated applications of PhTX-343 and spermine eventually resulted in inhibition. Higher concentrations of these compounds always inhibited the responses to acidic amino acids. Low concentrations of PhTX-343 and spermine also potentiated the responses to nicotine and gamma-aminobutyric acid. These results are discussed in terms of the postulated polyamine binding site on the N-methyl-D-aspartate receptor.

In the search for new toxins, preferably with new sites of action, the polyamine amides represent a new class of compounds with potential as insecticides and as pharmaceutical agents due to their antagonism of ligand-gated cation channels. In particular, they are potent antagonists of the l -glutamate receptors of insect skeletal muscle. In this paper, we report on synthetic studies to produce hybrid analogues based upon the argiotoxin spider toxins and philanthotoxin-433 which is obtained from a solitary, parasitic wasp. We speculate upon possible modes and sites of action for these antagonists and we discuss their potential as insecticides and in the possible treatment of ischaemic damage. The synthesis and characterization of 4-hydroxyphenylpropanoylspermine is reported and the locust muscle biological assay is described. Using this pharmacological screen, structure-activity relationships have been determined in our laboratories. These are reviewed in the light of the current literature. Voltage clamp studies of the synthetic analogue philanthotoxin-343 and the effects of this polyamine amide on glutamate receptors expressed in Xenopus oocytes are outlined. In conclusion, a description of our current ideas and understanding of the many sites and modes of action of the polyamine amides, based both upon our own studies and also upon those recently reported, is presented.