THE RETICULOSPINAL GLUTAMATE SYNAPSE IN LAMPREY - PLASTICITY AND PRESYNAPTIC VARIABILITY

1. The glutamatergic synapses formed between the unbranched giant reti culospinal axons onto spinal neurons in lamprey offer a central verteb rate synapse in which the presynaptic element can be impaled with one or several microelectrodes, which may be used for recording as well as microinjection of different substances. To provide a basis for the us e of this synapse in studies of release mechanisms, we have examined t he use-dependent modulation of the synaptic response under conditions of conventional cell body stimulation, and during direct stimulation o f the presynaptic axon. 2. To examine the stability of the mixed elect rotonic and chemical reticulospinal excitatory postsynaptic potential (EPSP) over time, action potentials were evoked at a rate of 1 Hz for 800-1000 trials. In three out of seven synapses the chemical component remained at a similar amplitude, while in four cases a progressive de crease (up to 35%) occurred. The electrotonic component remained at a similar amplitude in all cases. 3. During paired pulse stimulation of the reticulospinal cell body (pulse interval 65 ms) the chemical EPSP component showed a net facilitation in all cases tested [from 0.64 +/- 0.35 to 0.89 +/- 0.48 (SD) mV, n = 13], while the peak amplitude of t he electrotonic component was unchanged (1.37 +/- 0.68 and 1.36 +/- 0. 66 mV, respectively). Recording of the axonal action potential during paired pulse stimulation showed that the width of the first and second action potential did not differ [1/2 width (2.48 +/- 0.39 ms and 2.48 +/- 0.42 ms, respectively; n = 8)]. 4. The degree of facilitation var ied markedly between different synapses, ranging from an increase of a few percent to a two-fold increase (24 +/- 16% mean change of total E PSP amplitude, corresponding to 44 +/- 26% mean change of chemical EPS P amplitude). This type of variability was also observed in synapses m ade from the same unbranched reticulospinal axon onto different postsy naptic cells. 5. When paired pulse stimulation was applied to the reti culospinal axon in the very vicinity of the synaptic area (0.1-1 mm) a net depression of the chemical component occurred in 11 out of 19 cas es, and in the remaining cases the level of net facilitation was lower as compared with cell body stimulation (range between +17 and -23% ch ange of total EPSP amplitude; mean -5%; n = 19). 6. To test if the cha nge of the EPSP plasticity during local stimulation correlated with an increased transmitter release, two microelectrodes were placed in the same reticulospinal axon at different distances from the synaptic are a. The total EPSP amplitude was consistently larger, if the action pot ential was triggered with a depolarizing current pulse (1-2 ms) applie d close to the synapse (0.1-1 mm), than at a longer distance. During l ocal stimulation the EPSP amplitude increased with the magnitude of th e depolarizing stimulus pulse. If the axon was impaled closer than 0.1 mm from the synaptic area, however, the chemical EPSP became depresse d, but a reversal occurred after the electrode had been removed. 7. Th e increase of the total EPSP amplitude during local stimulation was ac companied by an increase of the peak amplitude of the initial electrot onic component, indicating that the amplitude of the presynaptic actio n potential was increased. Recording of the axonal action potential co nfirmed that both its amplitude and duration increased when it was tri ggered with depolarizing current pulses applied from an adjacent micro electrode. 8. To examine the anatomic correlate of a given reticulospi nal axon-motoneuron EPSP with an initial electrotonic component of 1.0 mV amplitude and a chemical component of 0.25 mV amplitude (distant s timulation), both elements were labeled intracellularly and a complete reconstruction of the synaptic connections was performed. Four axon-m otoneuron contacts, located on distal dendrites at 310-340 mu m distan ce from the motoneuron soma were observed, which altogether contained seven active zones, but only two gap junctions. Analysis of serial ult rathin sections showed that the different active zones were separated from each other by glial processes. 9. The present results show that t he use-dependent modulation of unitary glutamatergic EPSPs can vary be tween individual synapses along an unbranched axon, in which no activi ty-dependent changes in the shape or propagation of the presynaptic ac tion potential can be detected. If the stimulation electrode is placed within a distance of 1 mm from the synaptic area, however, the shape of the presynaptic spike, as well as the amplitude and use-dependent m odulation of the EPSP, can be markedly altered. These data should be t aken into account when large vertebrate axons are employed in studies of the mechanisms underlying synaptic glutamate release in the CNS.