Biophysical properties of the silent and activated rat sympathetic neuron following denervation.

A biophysical description of the denervated rat sympathetic neuron is reported, obtained by the two-electrode voltage-clamp technique in mature intact superior cervical ganglia in vitro. At membrane potential values negative to -50 mV, the normal, quiescent neuron displays voltage-dependent K and Cl conductances; following direct or synaptic stimulation (15Hz for 10 s), the neuron moves to a new resting state characterized by increased amplitude and voltage dependence of Cl conductance. Denervation produces two main effects: 1) resting Cl conductance gradually increases while its voltage-dependence decreases; by 30 days a high-conductance resting state prevails, almost independent of membrane potential in the -50/-110 mV range; 2) the increase in amplitude and voltage-dependence of Cl conductance, produced by direct stimulation in control neurons, is less marked in denervated neurons, and is observed over an increasingly small range of membrane potentials. Thirty days after denervation, the prevailing high-conductance resting state appears virtually insensitive to changes in membrane potential and stimulation. Voltage-dependent potassium currents involved in spike electrogenesis (the delayed compound potassium current and the fast transient potassium current) exhibit an early drastic decrease in peak amplitude in the denervated neuron; the effect is largely reversed after 6 days. Remarkable changes in fast transient potassium current kinetics occur following denervation: the steady-state inactivation curve shifts by up to +15 mV toward positive potential and voltage sensitivity of inactivation removal becomes more steep. A comprehensive mathematical model of the denervated neuron is presented that fits the neuron behavior under current-clamp conditions. It confirms that neuronal excitability is tuned by the conductances (mostly chloride conductance) that control the resting membrane potential level, and by fast transient potassium current. Impairment of the latter reduces both inward threshold charge for firing and spike repolarization rate, and fast transient potassium current failure cancels the voltage dependence of both processes.

Time course, localization and pharmacological modulation of immediate early inducible genes, brain-derived neurotrophic factor and trkB messenger RNAs in the rat brain following photochemical stroke.

A focal, unilateral thrombotic stroke was produced in the rat sensorimotor cortex. The time course of expression and localization of the immediate early inducible genes: c-fos, c-jun, zif268; nerve growth factor, brain-derived neurotrophic factor and the related tyrosine kinase high-affinity receptor (trkB) messenger RNAs were studied by in situ hybridization. The levels of messenger RNAs for c-fos, zif268, brain-derived neurotrophic factor (but not nerve growth factor) and trkB were consistently increased in cortex ipsilaterally to the lesion, while c-jun messenger RNA content was only slightly increased. The brain-derived neurotrophic factor messenger RNA was increased from 2 to 18 h following the stroke, mainly in cells having a normal morphological appearance. The trkB messenger RNA displayed temporal and spatial increases similar to brain-derived neurotrophic factor messenger RNA. The time course and pattern of expression of immediate early inducible gene and trophic factor messenger RNAs did not clearly support a causal relationship between these two families of factors. The observed messenger RNA increases were greatly attenuated by the non-competitive N-methyl-D-aspartate-sensitive glutamate receptor antagonist (+)-5-methyl-10,11-dihydroxy-5H-dibenzo(a,d)cyclohepten-5,10-imine , but substantially unaffected by the non-N-methyl-D-aspartate receptor antagonist 2,3-dihydroxy-6-nitrosulphanoylbenzoquinoxaline. The results suggest a major contribution of N-methyl-D-aspartate-sensitive glutamate receptor activation to the transcriptionally directed events subsequent to stroke. Future studies should clarify the contribution of these processes to either the progression of neuronal degeneration or the establishment of protective compensatory responses.

Nerve growth factor promotes functional recovery of retinal ganglion cells after ischemia.

PURPOSE: To investigate the effect of a transient complete ischemia on the function of cat retina and to determine whether nerve growth factor (NGF), which was previously shown to enhance retinal ganglion cell (RGC) survival after optic nerve section in the adult rat, can promote recovery of retinal neurons after the ischemic insult. METHODS: Function of distal and proximal retina was assessed by recording the electroretinogram in response to both homogeneous flickering light (FERG) and contrast reversing gratings (PERG), respectively, 30 days after the induction of a 60-minute episode of ischemia. Visual evoked potentials in response to contrast reversing gratings were also recorded to evaluate visual acuity and contrast thresholds. Cell survival after ischemia was assessed in retinal whole-mounts stained with cresyl violet. Cats were intraocularly treated with NGF every other day, 3 times a week, for 30 days. Controls were treated with either phosphate buffered saline or cytochrome c. RESULTS: After ischemia, the FERG was not significantly affected. On the contrary, the PERG, visual acuity, and contrast thresholds were severely impaired. After NGF treatment, PERG response amplitudes were much less reduced compared to controls, and visual acuity and contrast thresholds were virtually normal. In addition, a larger number of presumed RGCs was present in the NGF-treated retinas compared to the cyt c-treated ones. CONCLUSIONS: The more proximally located retinal neurons, in particular RGCs, are highly vulnerable to ischemia. Intraocular NGF treatment was effective in enhancing the survival and functional recovery of these neurons. This suggests that NGF may represent a novel therapeutic agent for the treatment of ischemic ocular pathologies.

Flash and pattern electroretinograms during and after acute intraocular pressure elevation in cats.

Retinal functionality during short-term intraocular pressure (IOP) elevation and simultaneous systemic blood pressure (BP) variations was evaluated in the cat by recording the electroretinogram in response to both homogenous flickering light (FERG) and contrast reversing gratings (PERG). The mean arterial blood pressure (BPm) was pharmacologically adjusted to different levels and a large range of IOP values was tested. Results indicate that both FERG and PERG responses are impaired when the eye perfusion pressure (PP = BPm - IOP) is reduced and they disappear at a critical PP value of about 20 mm Hg, irrespective of the absolute value of the IOP. In addition, when the critical perfusion pressure is maintained for periods longer than 5 min, the recovery of the PERG response, when present, is always delayed compared to the full recovery of the FERG response. These findings support the hypothesis that vascular factors, ie, the impairment of the retinal blood supply, may be responsible for the disappearance of the retinal electrical activity during short-term IOP elevation. Furthermore, the retinal ganglion cells, presumably the main source of the PERG response, appear less likely to recover from the acute ischemic episode.

Reproductive behavior of thalassemic couples segregating for Cooley anemia.

The reproductive behavior in 1984 of families segregating for Cooley anemia in Ferrara was compared with that of a control group of families, matched for some biological variables which affect fertility. At the resolution power of the sample, it was found that there is no significant difference in these variables due to segregation for Cooley anemia, and it appears that there is no longer significant reproductive compensation in thalassemic couples, although a tendency to compensate does still exist. The increased life span of children affected by Cooley anemia, due to improvements in treatment in the past decade, is probably the main reason why the compensatory reproductive behaviour of the past has almost disappeared.

Photochemical stroke and brain-derived neurotrophic factor (BDNF) mRNA expression.

In situ hybridization and Northern blotting were used to study the expression of brain-derived neurotrophic factor (BDNF) mRNA in the rat brain following photochemical stroke. A focal thrombotic lesion of the sensorimotor cortex was produced by intravenously injecting the light-sensitive dye rose bengal and exposing the skull to a controlled beam of light. Four hours after the light exposure the level of BDNF mRNA was increased in the hippocampus and cortex ipsilateral and perifocal to the lesion. The stroke-induced BDNF mRNA increase was prevented by the non-competitive glutamate receptor blocker dizocilpine (MK-801). The results indicate that the activation of N-methyl-D-aspartate (NMDA)-sensitive glutamate receptors is involved in the stroke-triggered stimulation of BDNF mRNA increase.

A model of signal processing at a mammalian sympathetic neurone.

A computational model has been developed for the action potential and, more generally, the electrical behaviour of the rat sympathetic neurone. The neurone is simulated as a complex system in which five voltage-dependent conductances (gNa, gCa, gKV, gA, gKCa), one Ca2+-dependent voltage-independent conductance (gAHP) and the activating synaptic conductance coexist. The individual currents are mathematically described, based on a systematic analysis obtained for the first time in a mature and intact mammalian neurone using two-electrode voltage-clamp experiments. The simulation initiates by setting the starting values of each variable and by evaluating the holding current required to maintain the imposed membrane potential level. It is then possible to simulate current injection to reproduce either the experimental direct stimulation of the neurone or the physiological activation by the synaptic current flow. The subthreshold behaviour and the spiking activity, even during long-lasting current application, can be analysed. At every time step, the program calculates the amplitude of the individual currents and the ensuing changes; it also takes into account the accompanying K+ accumulation process in the perineuronal space and changes in Ca2+ load. It is shown that the computed time course of membrane potential must be filtered, in order to reproduce the limited bandwidth of the recording instruments, if it is to be compared with experimental measurements under current-clamp conditions. The membrane potential trajectory and single current data are written in files readable by graphic software. Finally, a screen image is obtained which displays in separate graphs the membrane potential time course, the synaptic current and the six ionic current flows. The simulated action potentials are comparable to the experimental ones as concerns overshoot amplitude and rising and falling rates. Therefore, this program is potentially helpful in investigating many aspects of neurone behaviour.

N-(2-hydroxyethyl)hexadecanamide is orally active in reducing edema formation and inflammatory hyperalgesia by down-modulating mast cell activation.

Mast cells play a key role in inflammatory reactions triggered by tissue injury or immune perturbations. Little is known about endogenous molecules and mechanisms capable of modulating inappropriate mast cell activity. N-(2-Hydroxyethyl)hexadecanamide (palmitoylethanolamide), found in peripheral tissues, has been proposed to act as a local autacoid capable of negatively regulating mast cell activation and inflammation-hence the acronym Autacoid Local Inflammation Antagonism (ALIA). Recently, N-(2-hydroxyethyl)hexadecanamide (LG 2110/1) has been reported to down-modulate mast cell activation in vitro by behaving as an agonist at the peripheral cannabinoid CB2 receptor. Here, we have characterized and functionally correlated the anti-inflammatory actions of LG 2110/1 with its ability to control mast cell activation, when given orally in a battery of rodent models of inflammation. LG 2110/1 diminished, in a dose-dependent and correllated manner, the number of degranulated mast cells and plasma extravasation induced by substance P injection in the mouse ear pinna. In addition, LG 2110/1 reduced dose dependently plasma extravasation induced by passive cutaneous anaphylaxis reaction. In adult rats LG 2110/1 decreased, in a dose-dependent manner, carrageenan-induced hindpaw edema and hyperalgesia, but not phospholipase A2-induced hindpaw edema. Further, anti-edema effects were observed when utilizing dextran and formalin, known to also cause mast cell activation. Locally administered LG 2110/1 was likewise effective in minimizing dextran-induced hind paw edema. In contrast, equivalent amounts of palmitic acid plus ethanolamine were ineffective against plasma extravasation provoked by substance P. LG 2110/1 did not decrease plasma extravasation induced by the substance P fragment, substance P-(6-11), known to be inactive on mast cells. These results indicate that orally administered N-(2-hydroxyethyl)hexadecanamide is effective in: (a) directly down-modulating mast cell activation in vivo; (b) suppressing pathological consequences initiated by mast cell activation independently of the activating stimuli; (c) exerting an anti-inflammatory action distinguishable from that of classical steroidal and non-steroidal anti-inflammatory agents. These findings raise the possibility that N-(2-hydroxyethyl)hexadecanamide and related saturated N-acylamides ('ALIAmides') represent novel therapeutic agents useful in the management of inflammatory disease conditions.

The nicotinic activation of the denervated sympathetic neuron of the rat.

Nicotinic responses to endogenous acetylcholine and to exogenously applied agonists have been studied in the intact or denervated rat sympathetic neuron in vitro, by using the two-microelectrode voltage-clamp technique. Preganglionic denervation resulted in progressive decrease of the synaptic current (excitatory postsynaptic current, EPSC) amplitude, which disappeared within 24 h. These effects were accompanied by changes in ion selectivity of the nicotinic channel (nAChR). The extrapolated EPSC null potential (equilibrium potential for acetylcholine action, E(Syn)) shifted from a mean value of -15.9+/-0.7 mV, in control, to -7.4+/-1.6 mV, in denervated neurons, indicating a decrease of the permeability ratio for the main components of the synaptic current (P(K)/P(Na)) from 1.56 to 1.07. The overall properties of AChRs were investigated by applying dimethylphenylpiperazinium or cytisine and by examining the effects of endogenous ACh, diffusing within the ganglion after preganglionic tetanization in the presence of neostigmine. The null potentials of these macrocurrents (equilibrium potential for dimethylphenylpiperazinium action, E(DMPP); and equilibrium potential for diffusing acetylcholine, E(ACh), respectively) were evaluated by applying voltage ramps and from current-voltage plots. In normal neurons, E(Syn) (-15.9+/-0.7 mV) was significantly different from E(DMPP) (-26.1+/-1.0) and E(ACh) (-31.1+/-3.3); following denervation, nerve-evoked currents displayed marked shifts in their null potentials (E(Syn)=-7.4+/-1.6 mV), whereas the amplitude and null potential of the agonist-evoked macrocurrents were unaffected by denervation and its duration (E(DMPP)=-26.6+/-1.2 mV). It is suggested that two populations of nicotinic receptors, synaptic and extrasynaptic, are present on the neuron surface, and that only the synaptic type displays sensitivity to denervation.

Diversity of some gene frequencies in European and Asian populations. II. Fit of an isolation by distance model.

Six enzyme polymorphisms have been studied in European and Asian populations, using kinship as an index of genetic differentiation. Four clusters of populations are apparent, corresponding to four geographical regions. The differences between such groups account for a large fraction of genetic diversity, while minor differences are apparent between populations belonging to the same continent or subcontinent. The kinship as bioassayed from three loci (GLO, ESD, 6-PGD) correlates significantly with space, showing an exponential decline with the increase of distance between populations.

Functional and morphological abnormalities induced by ouabain intoxication of the rabbit retina.

Several researchers have recently used an intravitreal ouabain injection to induce a suitable model of experimental retinopathy and optic neuropathy in various animals. Ouabain administration into the vitreous body of rabbit causes an irreversible degeneration of the retinal layers and consequently of the optic nerve. The degeneration is proportional to the amount of injected drug. Electroretinographic recordings (ERG) show that these structural abnormalities are related to an inhibition of the electric retinal activity as the dose-dependent reduction of ERG waves amplitude has shown. Moreover, ERG and visual evoked responses (VER) measured at the same time evidence that the intravitreal injection of 1.7 nmol ouabain may block the impulse conduction along the optic nerve. This can be proved by the fact that 90 min after an ouabain injection VER disappears, while ERG is only partially reduced. These results are correlated with both morphological observation and autoradiographic studies on 3H-ouabain distribution in different retinal layers.

Can functional reorganization of area 17 following monocular deprivation be modified by GM1 internal ester treatment?

It has been extensively reported that monocular exposure early in life leads to profound alterations in visual cortical areas, where the majority of cells become responsive only to the stimulation of the normal eye. We have investigated a possible effect of the monosialoganglioside internal ester, termed AGF2, on the neuronal cortical plasticity of the kitten's visual cortex following monocular deprivation. Results indicate that in monocularly deprived kittens treated with ganglioside the ocular dominance shift in favor of the normal eye is partially prevented.

Synaptic stimulation of nicotinic receptors in rat sympathetic ganglia is followed by slow activation of postsynaptic potassium or chloride conductances.

Two slow currents have been described in rat sympathetic neurons during and after tetanization of the whole preganglionic input. Both effects are mediated by nicotinic receptors activated by native acetylcholine (ACh). A first current, indicated as IAHPsyn, is calcium dependent and voltage independent, and is consistent with an IAHP-type potassium current sustained by calcium ions accompanying the nicotinic synaptic current. The conductance activated by a standard synaptic train was approximately 3.6 nS per neuron; it was detected in isolation in 14 out of a 52-neuron sample. A novel current, IADPsyn, was described in 42/52 of the sample as a post-tetanic inward current, which increased in amplitude with increasing membrane potential negativity and exhibited a null-point close to the holding potential and the cell momentary chloride equilibrium potential. IADPsyn developed during synaptic stimulation and decayed thereafter according to a single exponential (mean tau = 148.5 ms) in 18 neurons or according to a two-exponential time course (tau = 51.8 and 364.9 ms, respectively) in 19 different neurons. The mean peak conductance activated was approximately 20 nS per neuron. IADPsyn was calcium independent, it was affected by internal and external chloride concentration, but was insensitive to specific blockers (anthracene-9-carboxylic acid, 9AC) of the chloride channels open in the resting neuron. It is suggested that gADPsyn represents a specific chloride conductance activatable by intense nicotinic stimulation; in some neurons it is even associated with single excitatory postsynaptic potentials (EPSCs). Both IAHP and IADPsyn are apparently devoted to reduce neuronal excitability during and after intense synaptic stimulation.

The slow Ca(2+)-activated K+ current, IAHP, in the rat sympathetic neurone.

1. Adult and intact sympathetic neurones of the rat superior cervical ganglion maintained in vitro at 37 degrees C were analysed using the two-electrode voltage-clamp technique in order to investigate the slow component of the Ca(2+)-dependent K+ current, IAHP. 2. The relationship between the after-hyperpolarization (AHP) conductance, gAHP, and estimated Ca2+ influx resulting from short-duration calcium currents evoked at various voltages proved to be linear over a wide range of injected Ca2+ charge. An inflow of about 1.7 x 10(7) Ca2+ ions was required before significant activation of gAHP occurred. After priming, the gAHP sensitivity was about 0.3 nS pC-1 of Ca2+ inward charge. 3. IAHP was repeatedly measured at different membrane potentials; its amplitude decreased linearly with membrane hyperpolarization and was mostly abolished close to the K+ reversal potential, EK (-93 mV). The monoexponential decay rate of IAHP was a linear function of total Ca2+ entry and was not significantly altered by membrane potential in the -40 to -80 mV range. 4. Voltage-clamp tracings of IAHP could be modelled as a difference between two exponentials with tau on approximately 5 ms and tau off = 50-250 ms. 5. Sympathetic neurones discharged only once at the onset of a long-lasting depolarizing step. If IAHP was selectively blocked by apamin or D-tubocurarine treatments, accommodation was abolished and an unusual repetitive firing appeared. 6. Summation of IAHP was demonstrated under voltage-clamp conditions when the depolarizing steps were repeated sufficiently close to one another. Under current-clamp conditions the threshold depolarizing charge for action potential discharge significantly increased with progressive pulse numbers in the train, suggesting that an opposing conductance was accumulating with repetitive firing. This frequency-dependent spike firing ability was eliminated by pharmacological inhibition of the slow IAHP. 7. The IAHP was significantly activated by a single action potential; it was turned on cumulatively by Ca2+ load during successive action potential discharge and acted to further limit cell excitability.

Monosialoganglioside (GM1) treatment of ouabain-induced retinopathy in the rabbit.

The mammalian retina is markedly influenced by cardiac glycosides. When nanomolar concentrations of ouabain are intravitreously injected into the rabbit, the visually evoked response completely disappears within 90 min, while scotopic ERG recordings show a remarkably decreased amplitude of the principal waves. When 33 nmol/kg monosialoganglioside are injected intravenously 30 min before topical intoxication, this functional impairment is significantly reduced. The electroretinographic response shows a limited amplitude reduction, while the cortical potential never disappears completely. Histological observations of intoxicated retinas show that a degenerative process begins in photoreceptor outer segment 24 h after the intravitreal ouabain injection. Presently, this process involves both the outer and inner nuclear layers and, finally, the ganglion cell layer. Comparing the intoxicated treated and untreated retinas, no difference is found in the degenerative pattern of the two groups. Autoradiographic studies are also reported to correlate the protective effect of monoganglioside (GM1) on this toxic retinopathy with its preferential accumulation in different retinal tissues.

Nicotinic EPSCs in intact rat ganglia feature depression except if evoked during intermittent postsynaptic depolarization.

The involvement of the postsynaptic membrane potential level in controlling synaptic strength at the ganglionic synapse was studied by recording nicotinic fast synaptic currents (EPSCs) from neurons in the intact, mature rat superior cervical ganglion, using the two-electrode voltage-clamp technique. EPSCs were evoked by 0.05-Hz supramaximal stimulation of the preganglionic sympathetic trunk over long periods; their peak amplitude (or synaptic charge transfer) over time appeared to depend on the potential level of the neuronal membrane where the nicotinic receptors are embedded. EPSC amplitude remained constant (n = 6) only if ACh was released within repeated depolarizing steps of the postganglionic neuron, which constantly varied between -50 and -20 mV in consecutive 10-mV steps, whereas it decreased progressively by 45% (n = 9) within 14 min when the sympathetic neuron was held at constant membrane potential. Synaptic channel activation, channel ionic permeation and depolarization of the membrane in which the nicotinic receptor is localized must occur simultaneously to maintain constant synaptic strength at the ganglionic synapse during low-rate stimulation (0.03-1 Hz). Different posttetanic (20 Hz for 10 s) behaviors were observed depending on the mode of previous stimulation. In the neuron maintained at constant holding potential during low-rate stimulation, the depressed EPSC showed posttetanic potentiation, recovering approximately 23% of the mean pretetanic values (n = 10). The maximum effect was immediate in 40% of the neurons tested and developed over a 3- to 6-min period in the others; thereafter potentiation vanished within 40 min of 0.05-Hz stimulation. In contrast, no statistically significant synaptic potentiation was observed when EPSC amplitudes were kept constant by repeated -50/-20-mV command cycles (n = 12). It is suggested that, under these conditions, posttetanic potentiation could represent an attempt at recovering the synaptic strength lost during inappropriate functioning of the ganglionic synapse.

Participation of a chloride conductance in the subthreshold behavior of the rat sympathetic neuron.

The presence of a novel voltage-dependent chloride current, active in the subthreshold range of membrane potential, was detected in the mature and intact rat sympathetic neuron in vitro by using the two-microelectrode voltage-clamp technique. Hyperpolarizing voltage steps applied to a neuron held at -40/-50 mV elicited inward currents, whose initial magnitude displayed a linear instantaneous current-voltage (I-V) relationship; afterward, the currents decayed exponentially with a single voltage-dependent time constant (63.5 s at -40 mV; 10.8 s at -130 mV). The cell input conductance decreased during the command step with the same time course as the current. On returning to the holding potential, the ensuing outward currents were accompanied by a slow increase in input conductance toward the initial values; the inward charge movement during the transient ON response (a mean of 76 nC in 8 neurons stepped from -50 to -90 mV) was completely balanced by outward charge displacement during the OFF response. The chloride movements accompanying voltage modifications were studied by estimating the chloride equilibrium potential (E(Cl)) at different holding potentials from the reversal of GABA evoked currents. [Cl(-)](i) was strongly affected by membrane potential, and at steady state it was systematically higher than expected from passive ion distribution. The transient current was blocked by substitution of isethionate for chloride and by Cl(-) channel blockers (9AC and DIDS). It proved insensitive to K(+) channel blockers, external Cd(2+), intracellular Ca(2+) chelators [bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA)] and reduction of [Na(+)](e). It is concluded that membrane potential shifts elicit a chloride current that reflects readjustment of [Cl(-)](i). The cell input conductance was measured over the -40/-120-mV voltage range, in control medium, and under conditions in which either the chloride or the potassium current was blocked. A mix of chloride, potassium, and leakage conductances was detected at all potentials. The leakage component was voltage independent and constant at approximately 14 nS. Conversely, gCl decreased with hyperpolarization (80 nS at -40 mV, undetectable below -110 mV), whereas gK displayed a maximum at -80 mV (55.3 nS). Thus the ratio gCl/gK continuously varied with membrane polarization (2.72 at -50 mV; 0.33 at -110 mV). These data were forced in a model of the three current components here described, which accurately simulates the behavior observed in the "resting" neuron during membrane migrations in the subthreshold potential range, thereby confirming that active K and Cl conductances contribute to the genesis of membrane potential and possibly to the control of neuronal excitability.

Synaptic current at the rat ganglionic synapse and its interactions with the neuronal voltage-dependent currents.

The membrane current activated by fast nicotinic excitation of intact and mature rat sympathetic neurons was studied at 37 degrees C, by using the two-microelectrode voltage-clamp technique. The excitatory postsynaptic current (EPSC) was modeled as the difference between two exponentials. A fast time constant (tau2; mean value 0.57 ms), which proves to be virtually voltage-independent, governs the current rise phase and a longer time constant (tau1; range 5.2-6.8 ms in 2 mM Ca2+) describes the current decay and shows a small negative voltage dependence. A mean peak synaptic conductance of 0.58 muS per neuron is measured after activation of the whole presynaptic input in 5 mM Ca2+ external solution (0.40 muS in 2 mM Ca2+). The miniature EPSCs also rise and decay with exponential time constants very similar to those of the compound EPSC recorded at the same voltage. A mean peak conductance of 4.04 nS is estimated for the unitary event. Deconvolution procedures were employed to decompose evoked macrocurrents. It is shown that under appropriate conditions the duration of the driving function describing quantal secretion can be reduced to <1 ms. The shape of the EPSC is accurately mimicked by a complete mathematical model of the sympathetic neuron incorporating the kinetic properties of five different voltage-dependent current types, which were characterized in a previous work. We show that IA channels are opened by depolarizing voltage steps or by synaptic potentials in the subthreshold voltage range, provided that the starting holding voltage is sufficiently negative to remove IA steady-state inactivation (less than -50 mV) and the voltage trajectories are sufficiently large to enter the IA activation range (greater than -65 mV). Under current-clamp conditions, this gives rise to an additional fast component in the early phase of membrane repolarization-in response to voltage pulses-and to a consistent distortion of the excitatory postsynaptic potential (EPSP) time course around its peak-in response to the synaptic signal. When the stimulation initiates an action potential, IA is shown to significantly increase the synaptic threshold conductance (up to a factor of 2 when IA is fully deinactivated), compared with that required when IA is omitted. The voltage dependence of this effect is consistent with the IA steady-state inactivation curve. It is concluded that IA, in addition to speeding up the spike repolarization process, also shunts the excitatory drive and delays or prevents the firing of the neuron action potential.

Development of diabetic neuropathy in the C57BL/Ks (db/db) mouse and its treatment with gangliosides.

We studied the development of diabetic neuropathy and its treatment with gangliosides using the C57BL/Ks mouse. The results of axonal morphometry showed the presence of a progressive axonal atrophy which was maximal at 180 days of age. To 400 days of age there was no longer any significant difference, perhaps due to aging processes. Nerve conduction velocity changed significantly from the early days of life. Thirty-day treatment with gangliosides significantly improved nerve conduction velocity and axonal morphometry at 180 and 280 days of life. No effect was observed with treatments at 30 or 60 days. It was previously shown that the early phase of the C57BL/Ks mouse neuropathy was reversed by insulin, whereas the late phase (180 days) was not. We showed elsewhere that at 180 days of age in the C57BL/Ks mouse there was a drastic decrease in slow transport of AChE (G1 and G2 molecular forms) indicating a shift in neuronal metabolism and suggesting that the disease was then more intrinsically neuronal. Using the suggestion of Robertson and Sima (Diabetes 29: 60-67, 1980) we label the first phase of the neuropathy "metabolic" (treatable with insulin) and the second phase "neuronal" (treatable with gangliosides). This "neuronal" phase could be related to the degenerative stage of human diabetic neuropathy.