Facilitation of transmitter release at squid synapses

Facilitation is shown to decay as a compound exponential with two time constants (T1, T2) at both giant and non-giant synapses in squid steilate ganglia bathed in solutions having low extracellular calcium concentrations ([Ca++]o). Maximum values of facilitation (F~) were significantly larger, and T1 was significantly smaller in giant than non-giant synapses. Decreases in [Ca++]o or increases in [Mn++]o had variable effects on T1 and F1, whereas decreases in temperature increased T~ but had insignificant effects on/'1. The growth of facilitation during short trains of equal interval stimuli was adequately predicted by the linear summation model developed by Mallart and Martin (1967.J. Physiol. (Lond.). 193: 676-694) for frog neuromuscular junctions. This result suggests that the underlying mechanisms of facilitation are similar in squid and other synapses which release many transmitter quanta.This work was supported by National Science Foundation research grant GB-36949, National Research Council (Canada) and Grass Fellowships to Dr. Charlton, and a National Institutes of Health career award (NS-00070) to Dr. Bittner.Neuroscienc

MA

thesisThis pilot descriptive study investigated the effects of a nursing procedure on selected cardiovascular hemodynamic variables. The nursing procedure consisted of turning and positioning the patients from a (1) supine to right lateral, (2) right lateral to supine, and (3) supine to left lateral position. Stroke volume, cardiac output, heart rate, peripheral resistance, systolic pressure and diastolic pressure were recorded during and at selected intervals after the nursing procedure. The computer-monitoring system measured these variables by obtaining measurements through a stiff Teflon catheter inserted into the aortic arch. The validity of this method has been previously established. Six consecutive adult patients with an aortic valve prosthesis were utilized from the patients brought to the Thoracic and cardiovascular Intensive Care Unit at the Latter-day Saints Hospital in Salt Lake City, Utah. Systolic and diastolic pressure demonstrated the widest changes and also the most frequent changes. Heart rate did not appear to directly influence cardiac output except in one patient. Decreases in arterial pressure appeared to be due to inadequate compensation in peripheral vascular resistance. Positioning the patient within 12 hours after surgery in the right lateral position appeared to affect the cardiovascular hemodynamic variables more than did positioning in the supine or left lateral position. The smallest clinically significant changes occurred when the patients were positioned in supine position which might suggest that if a patient were experiencing physiological instability, this would be the position of choice. The alterations of greatest significance in the cardiovascular hemodynamic variables occurred during the turning procedure. This would suggest the close observations of the patient for signs of impending shock such as changes in respiration, faintness and alterations in skin color and moisture, should be made during the turning and positioning procedure and not at a later time. Return of the variables toward the initial base line readings was almost instantaneous although complete restoration to the original readings varied with each patient. Limitations inherent in this study were the clinical setting, small sample, technical difficulties with the computer, and possible psychological effects of anxiety and fear. Even though the results are based a small sample, they are based on valid observations. If replication of this study supports the conclusion presented her, that systolic and diastolic pressure are altered significantly when positioned in the right lateral position, the nursing profession would be well advised to examine its policies regarding lateral positioning in the immediate post-operative period in patients with aortic valve replacements

Quantal Mechanisms Underlying Stimulation-induced Augmentation and Potentiation

Repetitive stimulation of motor nerves causes an increase in the number of packets of transmitter ( quanta ) that can be released in the ensuing period. This represents a type of conditioning, in which synaptic transmission may be enhanced by prior activity. Despite many studies of this phenomenon, there have been no investigations of the quantal mechanisms underlying these events, due to the rapid changes in transmitter output and the short time periods involved. To examine this problem, a method was developed in which estimates of the quantal release parameters could be obtained over very brief periods (3 s). Conventional microelectrode techniques were used to record miniature endplate potentials (MEPPs) from isolated frog (Rana pipiens) cutaneous pectoris muscles, before and after repetitive (40 sec at 80 Hz) nerve stimulation. Estimates were obtained of m (number of quanta released), n (number of functional release sites), p (mean probability of release) and var\rm\sb{s}p (spatial variance in p) using a method that employs counts of MEPPs per unit time. Fluctuations in the estimates were reduced using a moving bin technique (bin size = 3 s, $\Delta$bin = 1 s). Muscle contraction was prevented using low Ca\sp{2+}, high Mg\sp{2+} Ringer or normal Ringer to which $\mu$-conotoxin GIIIA was added. These studies showed that: (1) the post-stimulation increase in transmitter release was dependent on stimulation frequency and not on the total number of stimulus impulses. When the total number of pulses was kept constant, the high frequency pattern produced a higher level of transmitter release than did the lower frequency patterns; (2) augmentation and potentiation were present in both low Ca\sp{2+}, high Mg\sp{2+} and normal Ringer solutions, but potentiation, m, n, p and var\rm\sb{s}p were greater in normal Ringer solution than in low Ca\sp{2+}, high Mg\sp{2+} solution. In low Ca\sp{2+}, high Mg\sp{2+} solution, there was a larger decrease in n compared to p; (3) hypertonicity (addition of 100 mM sucrose) produced a marked increase in both basal and stimulation-induced values of m, n, and p. By contrast, there was a marked increase in the stimulation-induced but not the basal values of var\rm\sb{s}p; (4) hypertonicity produced a decrease in augmentation but had no effect on potentiation; (5) augmentation and potentiation appeared to involve mitochondrial uptake and efflux of cytoplasmic Ca\sp{2+}. Tetraphenylphosphonium (which blocks mitochondrial Ca\sp{2+} efflux and uptake) decreased augmentation and potentiation in low Ca\sp{2+}, high Mg\sp{2+} solutions but increased potentiation in the same solution made hypertonic with 100 mM sucrose; (6) the overall findings suggest that this new method may be useful for investigating the subcellular dynamics of transmitter release following nerve stimulation

Effects of high hydrostatic pressure on neuromuscular transmission in shallow-living and deep-living crustaceans

Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at the Massachusetts Institute of Technology and the Woods Hole Oceanographic Institution November, 1975The effects of high hydrostatic pressure on excitatory neuromuscular transmission in shallow- and deep-living crustaceans were compared. Pressure caused depression of the amplitude of excitatory junctional potentials (e.j.p.s) at the neuromuscular junction in the shallow-living crab, Libinia emarginata. A pressure of 100 atm depressed the e.j.p. amplitude by about one-half. In the deep- sea crab, Geryon quinquedens, which ranges to a depth of 2000 m (or 200 atm pressure), adaptations to high pressure were observed in two different types of muscle fibers: 1) In fibers with e.j.p.s that showed high levels of facilitation, the magnitude of pressure-induced depression decreased with increasing frequency of nerve stimulation; i. e., there was a pressure-induced increase in facilitation. Also a pressure-induced increase in the duration of the falling phase of the e.j.p. was observed which served to increase the level of depolarization resulting from summation of the e.j.p.s at high frequencies of nerve stimulation. In these highly facilitating fibers the physiologically significant frequencies that cause appreciable contraction are probably high. At high frequencies the pressure-induced increases in facilitation and summation together served to completely counteract the depressive effect of pressure, and the net depolarization attained during a train of nerve stimulation was relatively unaffected by pressures up to at least 200 atm. 2) Fibers with e.j.p.s showing low levels of facilitation may undergo significant contraction at low frequencies of nerve impulses where neither facilitation nor summation play a significant role. The amplitude of e.j.p.s recorded from this fiber-type in the deep-sea crab were, on the average, unaffected by pressures to 200 atm. The e.j.p.s of some of these fibers showed depression, but others were amplified under pressure. The results of experiments with the lobster, Homarus americanus, which ranges to a depth intermediate between Libinia and Geryon were in many respects intermediate between the results obtained with the two species of crab. Studies of the effect of pressure on isometric tension developed by whole muscles in Homarus and Geryon were consistent with the results of the studies of the e.j.p.; pressure depressed the rate of rise of tension in Homarus and had little effect in Geryon. The results of this work provides a physiological basis for the observation that shallow-living animals are generally immobilized by pressures in excess of 200 atm. Experiments were performed in an attempt to elucidate the mechanism underlying the pressure-induced depression of e.j.p. amplitude. Results were suggestive that the depression of e.j.p. amplitude reflects a pressure-induced decrease in the number of quanta of transmitter substance released by the nerve endings

PHYSIOLOGICAL AND ANATOMICAL ASSESSMENT OF SYNAPSES AT THE CRAYFISH NEUROMUSCULAR JUNCTION

The crayfish, Procambarus clarkii, has a multitude of ideal sites in which synaptic transmission may be studied. Its opener muscle, being innervated by a single excitatory neuron is a good model for studying the structure/function of neuromuscular junctions since the preparation is identifiable from animal to animal and the nerve terminals are visible using a vital dye. This allows ease in finding a suitable site to record from in each preparation and offers the ability to relocate it anatomically. Marking a recorded site and rebuilding it through electron microscopy gives good detail of synaptic struture for assesment.In the first of these studies, low output sites known as stems (which lie between varicosities) were used to reduce n (number of release sites) in order to minimize synaptic complexity so individual quantal events could be analyzed by their unique parameters (area, peak, tau, rise time and latency). This was in attempt to uncover specific quantal signatures that could be traced back to the structure of the area recorded. It was found that even at stem regions synaptic structure is still complex having multiple synapses each of which could harbor a number of AZs. This gives insight as to how quantal analysis should be treated. Even low output synapses n must be treated at the AZ level.Synaptic depression was studied at the crayfish extensor muscle. By depressing the phasic neuron and recording from the muscle it appears thatdepression is a presynaptic phenomenon. The use of 5-HT gave insight to vesicular dynamics within the nerve terminal, by delaying depression and increasing maximum EPSP amplitude. TEM of phasic nerve terminals reveals no change in numbers of dock or RRP vesicles. Short term facilitation and vesicular dynamics were studied with the use of 5-HT and a neurotoxin TBOA, which blocks the glutamate transporter. In this study I showed differential mechanisms that control RRP and RP vesicles. By blocking glutamate reuptake, the RRP is depleted as shown by reduced EPSPs, but recovered with 5-HT application. The understanding of vesicle dynamics in any system has relevance for all chemical synapses