The initial heat production in garfish olfactory nerve fibres.

A study has been made of the temperature changes associated with the passage of a single impulse in the non-myelinated fibres of the garfish olfactory nerve: and the time course of these temperature changes has been compared with the time course of the electrical events during the action potential. As in other non-myelinated nerves studied the observed temperature changes result from a biphasic initial heat production consisting of a transient evolution of heat (the positive heat) followed by a rapid heat reabsorption (referred to as the negative heat). There is no evidence of any additional phases of initial heat production. At 0 degrees C the measured positive initial heat is 224 mucal/g impulse (937 muJ/g impulse); and the corresponding negative initial heat is 230 mucal/g impulse (962 muJ/g impulse). The residual initial heat is very small, being about -6 mucal/g impulse (-25 muJ/g impulse). In the range 0-10 degrees C there is no significant effect of temperature on the magnitude of either the positive or the negative phases of heat production. The experimental thermal records were analysed to determine the true time course of the temperature changes in the nerve undistorted by the recording system. The time course of the temperature changes does not fit with that of the transmembrane voltage change as represented by the monophasic compound action potential recorded externally from the same point on the nerve. A better fit is obtained if the temperature changes are compared with the square of the voltage change in accordance with the view that the heat derives almost wholly from free energy changes and entropy changes in the membrane capacity. The best fit is obtained if it is assumed that the membrane potential does not discharge to zero during the action potential but that at the peak of the action potential the charge (and hence the p.d.) across the membrane capacity retains about 24% of its resting value.     

Heat production non-myelinated nerves.

Experiments with the C fibres of the rabbit vagus nerve have established that heat is evolved during the depolarizing phase of the action potential and is absorbed during the repolarizing phase. Subsequent studies using the pike olfactory nerve indicate that the heat production begins at a high rate very early in the depolarizing phase and is completed in advance of the peak of the spike. This would be expected if the heat arised from the energy released by the discharge of the membrane capacitance which varies as the square of the membrane potential; but estimates of the stored energy fall short of the observed heat production by a factor of two or three times. The prominent cooling phase suggests that a substantial part of the heat may arise from an entropy change. Such an entropy change would be expected to result from the change in the electrical stress in the dielectric of the membrane capacitance, and may thus be manifestation of reversible changes in the molecular architecture of the insulating matrix of the membrane.     

The effect of nerve activity on the distribution of synaptic vesicles

Certain gymnotid fish (apteronotids) continuously emit a high-frequency electric-organ discharge and thus continuously drive their electroreceptor afferents at high rates. Electroreceptor afferents terminate in one lamina of the electrosensory lateral line lobe (ELL) and can be readily sampled. Normally these terminals have many small vesicles clustered adjacent to the presynaptic membrane. When afferent activity is blocked for 24 hr by an injection of tetrodotoxin (TTX) into the electroreceptor nerve, the density of vesicles adjacent to the synaptic membrane declines; the volume of the remaining vesicles increases. If the nerve of a TTX-treated fish is stimulated proximal to the injection site, these changes can be reversed. These results imply that the migration of vesicles toward the presynaptic membrane is influenced by the level of activity in the nerve.     

Effect of amizil and arecoline on the activity of Na, K-ATP-ase and concentration of Na+ and K+ ions in rat brain]

A study was made of the activity of Na, K-ATP-ase and the Na+ and K+ content in the brain of rats with the action of arecoline and amizyl. Both arecoline and amizyl increased the Na, K-ATP-ase activity. This could be associated with the changes in the redistribution of the Na+ and K+ ions in the nerve cell. Arecoline proved to cause changes in the electrolyte distribution by the depolarization type, whereas amizyl--by the type of hyperpolarization of the nerve cell membrane.     

Effects of laser-induced hyperthermia treatment on ionic permeability of myelinated nerve

Summary The effect of laser-induced hyperthermia on the ionic permeability of nerve membranes was studied using the nodes of Ranvier in amphibian myelinated nerve as a model. To effect a photothermal modification of nerve membrane functions, con trolled laser irradiation consisting of a 5-sec thermal pulse was applied to the nodal membrane, increasing the temperature to a maximum of 48–58°C at the node. Major electrophysiological changes observed in the nodal membrane following laser-induced hyperthermia were a differential reduction of the sodium and potassium permeability, an increase in the leakage current, and a negative shift on the potential axis of the steady-state Na inactivation. There was no significant change in the kinetics of ion channel activation and inactivation for treatments below 56°C. The results suggest that a primary photothermal damage mecha nism at temperatures below 56°C could be a reduction in the number of active Na channels in the node, rather than a change in individual channel kinetics, or in the properties of the lipid bilayer of intervening nerve membrane. A differential heat sensi tivity between the noninactivated and the inactivated Na channels is also suggested. For the treatments of 56°C and above, a signifi cant increase of membrane leakage current suggests an irrevers ible thermal damage to the lipid bilayer. This work was supported by the ONR/SDIO N00014-86-K-0188 Medical Free-Electron-Laser Program and the Columbus-Cabrini Foundation.     

Millimeter wave effects on electrical responses of the sural nerve in vivo

Millimeter wave (MMW, 42.25 GHz)‐induced changes in electrical activity of the murine sural nerve were studied in vivo using external electrode recordings. MMW were applied to the receptive field of the sural nerve in the hind paw. We found two types of responses of the sural nerve to MMW exposure. First, MMW exposure at the incident power density ≥45 mW/cm² inhibited the spontaneous electrical activity. Exposure with lower intensities (10–30 mW/cm²) produced no detectable changes in the firing rate. Second, the nerve responded to the cessation of MMW exposure with a transient increase in the firing rate. The effect lasted 20–40 s. The threshold intensity for this effect was 160 mW/cm². Radiant heat exposure reproduced only the inhibitory effect of MMW but not the transient excitatory response. Depletion of mast cells by compound 48/80 eliminated the transient response of the nerve. It was suggested that the cold sensitive fibers were responsible for the inhibitory effect of MMW and radiant heat exposures. However, the receptors and mechanisms involved in inducing the transient response to MMW exposure are not clear. The hypothesis of mast cell involvement was discussed. Bioelectromagnetics 31:180–190, 2010. © 2009 Wiley‐Liss, Inc.     

The polypeptide and the phospholipid components of axon plasma membranes.

The axon plasma membrane fraction isolated from garfish olfactory nerve was analyzed for its polypeptide composition by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. There were present over 20 well-resolved polypeptide components in this membrane, and eleven of them, with an apparent molecular weight range of 22,000-130,000, accounted for most of the membrane proteins. None of the major polypeptide species present in the membrane appeared to be glycoprotein. Based on electrophoretic mobility on sodium dodecyl sulfate-polyacrylamide gel, eight of the major polypeptides found in garfish nerve membrane appeared to be also present in the axon plasma membrane isolated from lobster walking leg nerve. Both garfish and lobster nerve membranes contained high concentration of lipids (66-76%) which were essentially cholesterol and phospholipids. The classes of phospholipids present were phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, phosphatidylinositol and sphingomyelin. Lobster nerve membrane also contained about 3% phosphatidic acid. Assays for acetylcholinesterase in axon plasma membrane fractions isolated from different nerve sources showed a wide variation, ranging from a specific activity of 2.4 for garfish nerve to 312.5 for lobster nerve membrane.     

Effects of long-term heat exposure on the auditory nerve-brainstem evoked responses

1. 1.|The purpose of this study was to determine whether chronic latency changes were induced in the auditory nerve-brainstem potentials (ABR) during long-term heat exposure (acclimation).

2. 2.|Latency prolongations of the ABR were observed during acute (5 days) heat exposure. This was followed by a shortening of latencies and amplitude elevation after long-term (2 months) heat exposure (acclimation).

3. 3.|It was concluded that long-term exposure to heat induces chronic changes in nervous activity.

Rapid mechanical and thermal changes in the garfish olfactory nerve associated with a propagated impulse.

Mechanical and thermal changes associated with a propagated nerve impulse were determined using the garfish olfactory nerve. Production of an action potential was found to be accompanied by swelling of the nerve fibers. The swelling starts nearly at the onset of the action potential and reaches its peak at the peak of the action potential. There is a decrease in the length of the fibers while an impulse travels along the fibers. The time-course of the initial heat was determined at room temperature using heat-sensors with a response-time of 2-3 ms. Positive heat production was found to start and reach its peak nearly simultaneously with the action potential. The rise in temperature of the nerve was shown to be 23 (+/- 4) mu degrees C. In the range between 10 degrees and 20 degrees C, the temperature coefficient of heat production is negative, primarily due to prolongation of the period of positive heat production at low temperatures. The amount of heat absorbed during the negative phase varies widely between 45 and 85% of the heat evolved during the positive phase. It is suggested that both mechanical and thermal changes in the nerve fibers are associated with the release and re-binding of Ca-ions in the nerve associated with action potential production.     

Deficiency in matrix metalloproteinase gelatinase B (MMP-9) protects against retinal ganglion cell death after optic nerve ligation

Loss of retinal ganglion cells is the final end point in blinding diseases of the optic nerve such as glaucoma. To enable the use of mouse genetics to investigate mechanisms underlying ganglion cell loss, we adapted an experimental model of optic nerve ligation to the mouse and further characterized post-surgical outcome. We made the novel finding that apoptosis of retinal ganglion cells correlates with specific degradation of laminin from the underlying inner limiting membrane and an increase in gelatinolytic metalloproteinase activity. These changes co-localize with a specific increase in levels of the matrix metalloproteinase, gelatinase B (GelB; MMP-9). Using a transgenic mouse line harboring a reporter gene driven by the GelB promoter, we further show that increased GelB is controlled by activation of the GelB promoter. These findings led us to hypothesize that GelB activity plays a role in ganglion cell death and degradation of laminin. Applying the genetic approach, we demonstrate that GelB-deficient mice are protected against these pathological changes. This is the first report demonstrating a causal connection between GelB activity and pathological changes to the inner retina after optic nerve ligation.     

Kinetics of electron transport, proton transfer and photophosphorylation in chloroplasts and their relation to temperature-induced structural changes in the thylakoid membrane

Effects of various temperatures on the rates of electron transport between two photosystems, the light-induced uptake of protons, kinetics of proton efflux from the chloroplasts in the dark and photophosphorylation were studied in isolated chloroplasts. There are correlations between the physical state of thylakoid membrane and the rates of electron- and proton transport processes. The temperature dependence of "structural" parameter (fluidity of lipids in membrane) as well as the rates of electron- and proton transport processes reveal the breaks under the same temperatures. Stimulation of photophosphorylation by temperature increasing correlates with the heat activation of chloroplasts latent ATPase due to thermoinduced structural changes in the heat activation of chloroplasts latent ATPase due to thermoinduced structural changes in the protein part of CF0-CF1 complex. The rate of photophosphorylation also correlates with the physical state of membrane lipids. Thermoinduced "melting" of the thylakoid membrane inhibits the ATP formation because of a decrease in photosystem 2 photochemical activity and stimulation of membrane conductivity for protons.     

Na,K-atpase alterations in diabetic rats: relationship with lipid metabolism and nerve physiological parameters.

Type 1 diabetes induces several metabolic and biochemical disturbances which result in the alteration ofNa,K-ATPase, an enzyme implicated in the physiopathology of neuropathy Several fatty acid supplementations lessen this alteration. The aims of this study were to determine the possible relationships between Na,K-ATPase activity in nerves and red blood cells (RBCs) and, on one hand, the fatty acid alterations induced by diabetes in these tissues and plasma and on the other, on nerve physiological parameters. Two groups of rats, control and diabetic (n = 15), were sacrified 8 weeks after induction of diabetes with streptozotocin. Nerve conduction velocity (NCV), nerve blood flow (NBF), Na,K-ATPase activity and membrane fatty acid composition of sciatic nerves, red blood cells (RBCs) and plasma were measured. NCV, NBF and Na,K-ATPase activity in RBCs and in sciatic nerves were significantly decreased in diabetic rats. We revealed a positive correlation between Na,K-ATPase activity in sciatic nerves and both NBF and NCV and between Na,K-ATPase activity in RBCs and NBF and the same activity in sciatic nerve. Diabetes induced major changes in plasma fatty acids and RBC membranes and less important changes in sciatic nerve membranes. Na,K-ATPase activity correlated negatively with C20: 4 (n-6) and C22: 4 (n-6) levels in nerves and with C18: 2 (n-6) levels in RBCs. During diabetes, changes in the membrane fatty acid composition suggest the existence of a tissue-specific regulation, and the decrease in Na,K-ATPase activity correlates with the alteration in the level of specific fatty acids in RBCs and sciatic nerves. Modifications in the lipidic environment of Na,K-ATPase would be involved in the alteration of its activity. Na,K-ATPase activity seems to be implicated in the decrease of both NCV and NBF during diabetes.     

Properties of endogenous,membrane-associated sialidase activity (N-acetylneuraminidase) of the goldfish visual system

The endogenous sialidase (N-acetylneuraminidase) activity of membranes prepared from goldfish retina and optic tectum displays characteristics similar to those reported for neural plasma membrane sialidases of other organisms. Endogenous membrane sialidase activity was found to be optimal at ph 4.0, and maximal release was obtained at 37-50 degrees C, above which temperature thermal instability of the preparations was observed. Optic nerve crush, which results in regeneration of retinal ganglion cell axons, did not result in significant changes in measured endogenous membrane sialidase activity in either the retina or the optic tectum. Enzymatic hydrolysis of membrane sialoglycolipid (ganglioside) accounted for about 70% of the total sialic acid released. Ganglioside GM1 accumulated as the major lipid product in both retina and tectum, indicating that the inner sialosylgalactosyl linkage in the ganglio oligosaccharide series was resistant to hydrolysis by the endogenous enzyme.     

Heat shock-induced changes in lipid and protein metabolism in the endoplasmic reticulum of barley aleurone layers

Heat shock in barley aleurone layers induces heat shock protein synthesis and suppresses secretory protein synthesis by selectively destabilizing their mRNAs. In addition, the endoplasmic reticulum (ER) membranes upon which secretory protein mRNAs are translated become vesiculated during heat shock, leading to the hypothesis that ER dissociation and targeted mRNA destabilization are linked mechanistically. Supporting this, ER can be heat adapted, and heat-adapted ER has higher levels of fatty acid saturation in membrane phospholipids which do not vesiculate upon heat shock. Secretory protein mRNAs are also more stable in heat-adapted cells. To understand better heat shock-induced changes in ER membranes, we examined ER membrane proteins and enzymes involved in phosphatidylcholine biosynthesis and phospholipid turnover in heat-shocked aleurone cells. Heat shock significantly increased the activity of phospholipases A2 and D, and shortly thereafter significant but gradual increases in choline kinase and phosphocholine glyceride transferase activities and a sharp increase in phosphorylcholine citidyl transferase activity were observed. Only minor changes were observed in SDS-PAGE analyses of proteins from sonicated ER membranes fractionated on continuous sucrose gradients. Overall, heat shock reduced total lipid in ER membranes relative to protein, and in intact, ultracentrifuged aleurone cells examined by light and electron microscopy the ER band appeared to increase in density. The changes in phospholipid metabolism coupled with the suppression of secretory protein synthesis indicate that in addition to inducing a classic heat shock response, high temperature also induces a classic unfolded protein response in the ER of this secretory cell.     

Activity of Photosystem 2, Lipid Peroxidation,and the Enzymatic Antioxidant Protective System in Heat Shocked Barley Seedlings

The impact of heat shock on minimising the activity of photosystem 2 (PS2) initiating high lipid peroxidation (POL) level and consequently changes in the enzymatic-antioxidant protective system was studied in seedlings of two Egyptian cultivars of barley (Giza 124 and 125). Heat doses (35 and 45 °C for 2, 4, 6, and 8 h) decreased chlorophyll (Chl) contents coupled with an increase in Chl a/b ratio, diminished Hill reaction activity, and quenched Chl a fluorescence emission spectra. These parameters reflect the disturbance of the structure, composition, and function of the photosynthetic apparatus as well as the activity of PS2. POL level, as dependent on the balance between pro- and anti-oxidant systems, was directly correlated with temperature, exposure time, and their interaction. Heat shock caused an increase in the electric conductivity of cell membrane, and malonyldialdehyde content (a peroxidation product) coupled with the disappearance of the polyunsaturated linolenic acid (C_18:3), reflecting the peroxidation of membrane lipids which led to the loss of membrane selective permeability. Moreover, it induced distinct and significant changes in activities of antioxidant enzymes. Superoxide dismutase and peroxidase activities have been progressively enhanced by moderate and elevated heat doses, but the most elevated one (45 °C for 8 h) showed a decrease in activities of both enzymes. In contrast, catalase activity was reduced with all heat shocks.     

The effect of anaesthetics on protein conformation in membranes as studied by the spin-labelling technique

The effect of general and local anaesthetics was studied on the membrane components (proteins and lipids) of frog sciatic nerve using the spin-labelling technique. A rapid and effective method was developed to incorporate fatty acid labels into the lipid regions and to attach the maleimide spin labels to the thiol sites of the proteins of the membrane. On the basis of the rotational correlation time of the attached maleimide spin labels, at least three different protein domains were identified. One part of the labels has a preferred orientation with respect to the axis of the nerve fibre. The effect of halothane and of local anaesthetics such as lidocaine and tetracaine, which influence primarily the lipid regions of the membrane, is efficiently transferred to the spin-labelled membrane proteins via strong lipid-protein interaction. The results support the concept that the architecture and the physiological activity of the membrane-bound proteins are sensitive to changes in the physical state of membrane lipids.     

Effects of heating and cooling on nerve terminal impulses recorded from cold-sensitive receptors in the guinea-pig cornea

An in vitro preparation of the guinea-pig cornea was used to study the effects of changing temperature on nerve terminal impulses recorded extracellularly from cold-sensitive receptors. At a stable holding temperature (31-32.5 degrees C), cold receptors had an ongoing periodic discharge of nerve terminal impulses. This activity decreased or ceased with heating and increased with cooling. Reducing the rate of temperature change reduced the respective effects of heating and cooling on nerve terminal impulse frequency. In addition to changes in the frequency of activity, nerve terminal impulse shape also changed with heating and cooling. At the same ambient temperature, nerve terminal impulses were larger in amplitude and faster in time course during heating than those recorded during cooling. The magnitude of these effects of heating and cooling on nerve terminal impulse shape was reduced if the rate of temperature change was slowed. At 29, 31.5, and 35 degrees C, a train of 50 electrical stimuli delivered to the ciliary nerves at 10-40 Hz produced a progressive increase in the amplitude of successive nerve terminal impulses evoked during the train. Therefore, it is unlikely that the reduction in nerve terminal impulse amplitude observed during cooling is due to the activity-dependent changes in the nerve terminal produced by the concomitant increase in impulse frequency. Instead, the differences in nerve terminal impulse shape observed at the same ambient temperature during heating and cooling may reflect changes in the membrane potential of the nerve terminal associated with thermal transduction.     

Membrane-potential-sensitive dyes for optical monitoring of activity in Aplysia neurons

Two membrane-associated dyes (WW375 and NK2367) which change their absorption of light when the membrane potential changes have been studied using several preparations from Aplysia. Action potentials are easily observed in nerve trunks (from a number of axons), in bag cell clusters, in some of the larger single cells of the parietovisceral ganglion, and in the optic nerve. Physiological effects of the dyes on the circadian rhythm of activity in the eye are described.     

Changes in Infrared Spectrum of Nerve during Excitation

MOLECULAR changes that occur in the membrane of nerve during depolarization are important to explain the mechanism of excitation and conduction. During an action potential in nerve the electrical field strength across the membrane undergoes a change of about 10⁵ V cm^?1 which could influence the conformation of membrane molecules possessing a flexible dipole¹. Several methods have been reported for detecting changes with excitation in nerve, muscle and electroplaque^2–11, but have been difficult to interpret. We have therefore used an adaptation of the method of modulation spectroscopy¹², which is based on changes in the absorption spectrum as a result of perturbing the molecular structure^13–16. Our method was based on absorption in the infrared, which occurs for molecular vibrations accompanied by changes in dipole moment.     

The plasma membrane of yeast acquires a novel heat-shock protein (hsp30) and displays a decline in proton-pumping ATPase levels in response to both heat shock and the entry to stationary phase.

Recent studies have revealed that the action of the proton-translocating ATPase of the plasma membrane of yeast is an important determinant of several stress tolerances and affects the capacity of cells to synthesise heat shock proteins in response to heat shock [Panaretou, B. & Piper, P. W. (1990) J. Gen. Microbiol. 136, 1763-1770; Coote, P. J., Cole, M. B. & Jones, M. V. (1991) J. Gen. Microbiol. 137, 1701-1708]. This study investigated the changes to the protein composition of the Saccharomyces cerevisiae plasma membrane that result from a heat shock to dividing cultures and the entry to stationary growth caused by carbon source limitation. Plasma membranes were prepared from exponential, heat-shocked and stationary yeast cultures. The proteins of these membrane preparations were then analysed by polyacrylamide gel electrophoresis and immunoblot measurement of ATPase levels. The protein composition of plasma membranes displayed two prominent changes in response to both heat shock and the entry to stationary phase: (a) a reduction in the level of the plasma membrane ATPase; and (b) the acquisition of a previously uncharacterised 30 kDa heat-shock protein (hsp30). The ATPase decline with heat shock probably exerts an important influence over the ability of the cell to maintain ATPase activity, and therefore intracellular pH, during extended periods of stress. Through in vivo pulse-labelling of plasma membrane proteins synthesised before and during heat shock, followed by subcellular fractionation, it was shown that hsp30 is the only protein induced by the yeast heat-shock response that substantially copurifies with plasma membranes. It might therefore exert a stress-protective function specifically at this membrane.