Modulation of synaptic transmitter release by repetitive postsynaptic action potentials.

The effect of repetitive action potentials in the postsynaptic axon on the release of synaptic transmitter from the presynaptic terminal was investigated at the squid giant synapse. Repetitive antidromic stimulation of the postsynaptic axon resulted in a reduction in the excitatory postsynaptic potential (EPSP). The reduction in transmitter release was accompanied by a decrease in the presynaptic spike after-hyperpolarization (AH). Increasing the concentration of extracellular potassium ions also reduced the EPSP and decreased the amplitude of the presynaptic spike AH. The reduction in transmitter release resulting from repetitive postsynaptic impulses is attributed to the accumulation of extracellular potassium ions. It is proposed that the accumulation of extracellular potassium ions resulting from repetitive postsynaptic activity may modulate synaptic transmission and function as an integrative mechanism in the nervous system.

Centenary of the synapse: from Sherrington to the molecular biology of the synapse and beyond.

Few concepts have meant more to neuroscience than the synapse, commonly understood to mean the junction between two excitable cells. The term was introduced by Charles Sherrington in 1897. The centenary of this event is an appropriate time to review the term's origins and utility. There are some surprises. The term didn't actually come from him. His concept was more functional than structural. The pioneering physiological and structural studies in the 1950s in fact did not lead to a rigorous definition. There is still confusion on how to define neurotransmitters. As molecular biological approaches are increasingly refining the concept of a fundamental synaptic unit, many types of neuronal interactions are appearing that do not fit with the synaptic concept. Are the neural circuits underlying behaviour strictly synaptic? In dealing with these questions, a longer perspective is useful for understanding how the term arose, how it has evolved to the present, and what kinds of challenges may be coming in the future.

Interactions among lumbar motoneurons on opposite sides of the frog spinal cord: morphological and electrophysiological studies.

Light and electron microscopy have been used to study the projections of dendrites from motoneurons in lumbar segments of the spinal cord of the frog following administration of horseradish peroxidase to cut ventral roots. Processes originating from motoneurons crossed to the opposite side of the spinal cord via the anterior commissure and made contact with dendrites and motoneuronal somata. Typically, in segments 6 to 8 the crossing dendrites showed irregular enlargements in diameter. Electrophysiological recordings were obtained both extracellularly from ventral roots and intracellularly from motoneuronal somata. In Ringer's solution containing 1 mM calcium, stimulation of a lumbar ventral root, elicited population responses with early and late components in the ventral root of the opposite side of the same segment. Only the early, short latency component remained in calcium-deficient Ringer's solution. In calcium-containing Ringer's solution, intracellular recording from an antidromically activated motoneuron showed an action potential with a short latency; this response was followed by excitatory postsynaptic potentials (epsps) from which action potentials could be generated. Contralateral ventral root stimulation also elicited in the same motoneuron a short latency action potential that was rarely followed by epsps. The short latency responses, that were elicited by stimulation of ventral roots of either side persisted in calcium-deficient Ringer's solution, but the epsps were abolished. Contralaterally elicited short latency responses were eliminated by section of the anterior commissure. We believe that electrically mediated crossed interactions among lumbar motoneurons may serve as a means of coordinating muscle groups of opposite sides that are used in movements that require bilateral synchronization, such as jumping and swimming.

Macrophages direct process elongation from adult frog motorneurons in culture.

Motorneurons and macrophages have been isolated and identified in primary cultures from adult frog (Rana pipiens) spinal cord. Time-lapse video microscopy revealed that during the first two weeks migrating macrophages contact the growth cones of motorneurons. As they continue to migrate, the motorneuron processes elongate in close association with the moving macrophages. Elongating motorneuron processes are thereby brought into contact with other motorneurons and networks are formed. At later stages, the macrophages die but the motorneurons and the networks survive for at least another two weeks. These experiments show that macrophages can promote a directed elongation of motorneuron processes and suggest that they play a similar role during regeneration in vivo.

The influence of 17 beta-oestradiol on K+ currents in smooth muscle cells isolated from immature rat uterus.

Potassium currents in cells isolated from myometrium of immature (23-26 days after birth) rats were measured by whole-cell patch clamp to analyse the effects of gonadal steroids. Predominantly outward K+ currents with an early transient component were recorded in response to depolarizing pulses from a holding potential of -90 mV. The transient current, which was rarely present in adult myometrial cells, was inactivated by holding the membrane potential at -40 mV and attenuated by 1 mM 4-aminopyridine. Under these conditions a second sustained outward current was apparent. Administration of 17 beta-oestradiol to the immature rat before isolation of the myometrial cells reduced the probability of occurrence of the transient currents from 78.6% of cells (n = 33) to 29.8% (n = 14). Progesterone had only a slight effect. The kinetics of the transient currents were differently influenced by the two steroids. Cells isolated from animals previously given 17 beta-oestradiol showed transient outward currents with significantly shorter time constants of decay than those for control cells. Conversely, progesterone caused the time constants to be increased. We conclude that the gonadal steroids exert an influence on the expression of different populations of ionic channels in isolated cells of the immature rat uterus. These channels may regulate the excitability and contractility of the uterus in vivo.

Intracellular and extracellular calcium ions in transmitter release at the neuromuscular synapse.

The theme of this presentation has been to show that the control of transmitter release at the neuromuscular synapse is achieved by extracellular and intracellular calcium. For the fast information transfer represented by the end-plate potential, the electrochemical gradient for calcium across the presynaptic membrane and the associated calcium conductance seem to play the primary role. For slower processes such as tetanic and posttetanic potentiation, the combined effect of both sources for calcium determine the amount of transmitter liberated.

Hormonal influence on ionic channels in myometrium.

Smooth muscle cells in culture isolated from myometrium were characterized by scanning microscope and immunohistochemistry. Using the whole-cell patch-clamp configuration, and the single channel bilayer technique, the properties of ionic channels expressed in both non-pregnant and pregnant myometrium have been described. The predominantly expressed potassium channel changes from a transient inactivating outward current seen before puberty, to a calcium sensitive delayed outward current present in the adult stage. A change in the calcium channel population occurs from the nonpregnant to the pregnant state. Finally, sodium channels are expressed with greater frequency towards the end of gestation suggesting that these channels may play a role in labor.

The bulbo-spinal indoleaminergic pathway in the frog.

The role of the bulbo-spinal indoleaminergic pathway of the frog was investigated. Using the isolated spinal cord preparation, responses of motoneurons of segments 9 and 10 to lateral column stimulation were recorded from ventral roots (LC-VRP) and by intracellularly placed microelectrodes. Control responses were compared to those obtained after addition of substances known to alter indoleaminergic synthesis or receptor activation. Responses from spinal cords of animals that were pretreated with indoleamine-depleting agents were compared to mean control responses. Procedures that inhibited indoleamine synthesis or blocked indoleamine receptors reduced motoneuronal activity. This was manifested as an increase in mean latency and a decrease in amplitude of the monosynaptic LC-VRP, an increased duration of suppression following an LC conditioning stimulus, and a decrease in spontaneous activity. In contrast to intracellularly recorded responses from control cords, those recorded under these conditions typically showed single spikes with longer latencies. Additions of indoleamine precursors to normal cords or to cords depleted of monoamines by reserpine shortened mean latencies, increased amplitudes of LC-elicited responses and caused an increase in spontaneous activity. These observations were recorded both extracellularly and intracellularly. Our results suggest that the bulbo-spinal indoleaminergic pathway modulates the output of motoneurons of the frog spinal cord.

A hypothesis for the superior sensitivity of wheat germ agglutinin as a neuroanatomical probe.

We have shown that a conjugate of wheat germ agglutinin (WGA), with horseradish peroxidase (HRP), is more sensitive than native HRP as a probe of neuroanatomic connections involving the retrograde transport of the lectin. It has also been shown in our laboratory that WGA-HRP remains at the site of injection twice as long as HRP. The purpose of the present morphometric study was to investigate the basis for the higher sensitivity of WGA-HRP over HRP as a retrogradely transported tracer molecule. To do this, we modified the experiment of Heuser and Reese which utilized the tracing of HRP in the frog neuromuscular junction (Heuser, J.E. and Reese, T.S., J. Cell Biol., 57 (1973) 315-344). Instead of using HRP alone, we examined, in double labeling experiments, fluid and adsorptive endocytosis with free HRP and WGA coupled to ferritin (WGA-ferritin) respectively. Immediately after nerve stimulation, both markers are taken up simultaneously into cisternae, and in tubular structures strikingly similar to the described compartment of uncoupling of receptor from ligand (CURL). Frequently, cisternae were connected with putative CURL. This early double labeling of cisternae and putative CURL was followed by the appearance of synaptic vesicles labeled with WGA-ferritin only (72-79%), HRP only (6-11%), and both labels (13-16%). In contrast to the labeling pattern of synaptic vesicles, the majority of cisternae and putative CURL had both labels throughout the duration of the experiments (77-80%). The results of this study indicate that most of WGA-ferritin and HRP are co-localized in cisternae and putative CURL, compartments involved in endocytosis and surface receptor recycling.(ABSTRACT TRUNCATED AT 250 WORDS)

Dye and electrical coupling between frog motoneurons.

Coupling between lumbar motoneurons in the isolated frog spinal cord was studied by using intracellular recordings and intracellular injections of Lucifer Yellow CH. Physiological studies revealed intra- and intersegmental, short latency electrical interactions between many motoneurons. Injections of Lucifer Yellow into motoneurons that were found to be coupled electrically revealed intra- and intersegmental dye-coupling.

The action of the sodium ionophore, monensin, or transmitter release at the frog neuromuscular junction.

The action of the sodium ionophore, monensin, on spontaneous and evoked transmitter release at the frog neuromuscular junction was studied. Ringer's solutions with low calcium concentrations (0.4 mM or 0 Ca 1 mM EGTA) were used to bathe the preparation. Following addition of monensin in the bathing solution: (1) substantial increases in miniature end-plate potential frequency occurred; (2) tetanic nerve stimulation caused increases in potentiation of approximately 10 times over control values; (3) there was a dual action on the amplitude of the end-plate potential. We conclude that sodium ions take part in the regulation of transmitter release at the neuromuscular junction.

The role of calcium ions in tetanic and post-tetanic increase of miniature end-plate potential frequency.

1. The role of Ca ions in transmitter release changes, during and after high frequency stimulation of the motor nerve (10--100 Hz), was examined at the frog neuromuscular junction. 2. The stimulation-induced changes in miniature end-plate potential frequency (f) resembled the changes in end-plate potential amplitude recently described by Magleby and Zengel (1975, 1976). 3. The effects of tetanic stimulation on f under inward electrochemical gradient for Ca ions were compared with those under reversed gradient and four differences were found: (a) The increase in f during the tetanus under reversed Ca gradient conditions is much smaller than with an inward Ca gradient. (b) The increase in f under reversed Ca gradient is preceded by a small decrease in f, whereas with an inward Ca gradient an immediate increase in f is observed. (c) After the termination of the tetanus with a reversed Ca gradient, there is a further increase in f, compared to a decrease with an inward Ca gradient. (d) The augmentation phase of post-tetanic potentiation was practically abolished. 4. The experimental results are explained by assuming that high frequency nerve stimulation causes an increase in transmitter release by at least two distinct processes: influx of Ca ions through the presynaptic membrane and release of Ca ions from intracellular stores. It is suggested that Na ions couple nerve activity to intracellular release of Ca.

The action of calcium at spinal neurones of the frog.

1. The action of Ca on membrane excitability and synaptic transmission at motoneurones of the isolated spinal cord of the frog has been studied.2. Impulse propagation along presynaptic fibres was unaffected by [Ca] in the range of 0-10 mM in the presence of 1 mM-Mg.3. Motoneurone membrane excitability was within normal ranges when the cord was bathed in low or 0 Ca solutions containing 1 mM-Mg but was depressed in 5 or 10 mM-Ca solutions.4. Spontaneous miniature synaptic potentials were recorded in the presence of tetrodotoxin. The frequencies were reduced but not abolished in the absence of bath Ca, but were increased greater than twofold in 5 or 10 mM-Ca in comparison to those in normal Ringer solution (1 mM-Ca).5. Iontophoretic application of Ca both near to and remotely from the intracellular recording site (presumably in the soma) caused facilitation of elicited post-synaptic potentials.6. It is concluded that the action of Ca at these central synapses is identical to that described at the neuromuscular junction.