344 DAVID R. CURTIS 



barrel, the orifice of the outer barrel remaining in the extracellular position, 

 so that chemical substances can be applied into this extracellular space 

 iontophoretically (cf. Curtis and Eccles, 1958; Curtis et al., 1960a; Curtis and 

 Watkins, 1960b). The use of such an electrode therefore enables the resting 

 potential of the impaled cell to be measured, the membrane conductance to 

 be evaluated and the presence and magnitude of postsynaptic and spike 

 potentials to be determined. These studies can be performed before, during 

 and after the application of the chemical agent and if a hyperpolarization is 

 so produced, the similarity between the responsible membrane permeabiHty 

 alteration and that producing the i.p.s.p. can be assessed. The membrane 

 potential of the cell can be varied by applying current through the intra- 

 cellular electrode and also the ionic concentrations of the intracellular 

 medium can be modified by iontophoretic injections of ions from this 

 electrode. Such techniques established that the equihbrium potential of the 

 ions that diffuse across the receptor membrane to produce the i.p.s.p. is 

 approximately 10 mV more hyperpolarized than the resting potential of 

 motoneurons (— 70mV) and that the responsible ions are probably K+ and 

 Cl~ (Coombs et al., 1955). If the equilibrium potential of the change in 

 membrane potential produced by the applied chemical agent is the same as 

 that of the i.p.s.p., the possibility that it is a transmitter would be very strong 

 and would be further strengthened by the finding that the membrane con- 

 ductance of K+ and Cl~ ions was also increased. 



It must be assumed that excitatory and inhibitory transmitter substances 

 are not identical (Grundfest, 1957) and that the permeabihty changes at the 

 respective synapses are specific to the type of synapse not only because of 

 the type of receptor involved but also because of the nature of the chemical 

 transmitter liberated. If this were not so, diffuse apphcation of a transmitter 

 to the neuron, although producing an increase in membrane conductance, 

 may fail to change the membrane potential if approximately equal numbers 

 of excitatory and inhibitory synapses were activated. It would then be 

 necessary to apply the substance to the immediate environment of the 

 subsynaptic receptors of one synapse. Such a technique is clearly impossible 

 and is rendered unnecessary by the evidence to be discussed below, that 

 excitatory and inhibitory transmitters, at least in the spinal cord, are different 

 substances. 



Several substances which have been investigated by these methods have 

 failed to alter the resting membrane potential of motoneurons although the 

 excitability of these cells has been modified. Thus y-amino-«-butyric acid, 

 j8-alanine and some related monocarboxylic amino acids (Curtis et al., 1959; 

 Curtis and Watkins, 1960b), although increasing the membrane conductance 

 of motoneurons, fail to produce a membrane hyperpolarization and have 

 therefore been rejected as possible inhibitory transmitters. The apphcation of 

 a high concentration of certain dicarboxylic amino acids also depresses 



