Postsynaptic Axon Research Articles

F58. Separation of human central pain and temperature pathways

Facial sensations of sharp (pin), warm, cool and touch are transmitted by different peripheral fibers. Animal studies show separate trigeminal nucleus caudalis (NC) neurons responding to pain and temperature modalities but also possibly some polymodal neurons. Traditionally, the animal and human central sensory pathways are described as follows. Pain and temperature fibers travel together to synapse in the inferior trigeminal spinal nucleus, NC. Post-synaptic axons cross the midline in a single pathway to form the pain and temperature trigemino-thalamic pathway. Larger myelinated touch fibers synapse in the brainstem 5th nucleus but smaller touch fibers synapse in spinal nuclei. Disruption of crossing fibers at the midline causes simultaneous pain and temperature deficits in syringomyelia. This report describes 11 syringomyelia patients whose deficits provide evidence for separate, central afferent, post-synaptic pathways for facial pain, warm and cool sensations. Evaluations: MRIs and exams including stimuli at 21–22 °C and 40–41 °C. Exams show discrepancies between warm and cool sensations and between pin and temperature sensations. Preserved touch is present in 3 patients with pin deficit. Three cases lack MRI abnormalities appropriate to their deficit, but 2 of those probably had syringes that were previously larger. Translation of experimental animal studies to human function benefits from verification. Unlike human case reports and animal studies, the spinal cord lesions in our patients did not cause symmetrical pain and temperature deficits. Most human reports of temperature sensation don’t indicate what thermal stimulus was used. Use of careful warm and cool stimuli would likely find additional discrepant cases that also suggest distinct but probably adjacent pain, warm and cool central sensory pathways. Differences between touch and pin sensation may be due to the bilateral touch pathways.

Critical role for Annexin A7 in secondary brain injury mediated by its phosphorylation after experimental intracerebral hemorrhage in rats

Glutamate excitotoxicity has been implicated in intracerebral hemorrhage (ICH)-induced secondary brain injury (SBI). Synaptosome associated protein 23 (SNAP23) and SNAP25 are respectively participate in presynaptic glutamate release and postsynaptic glutamate receptor (NMDA receptor) trafficking, both of which are essential for glutamate-mediated excitatory toxicity. SNAP23 and SNAP25 exhibit high homology and SNAP23 has been shown to interact with Annexin A7 (ANXA7). This study was to examine the role of ANXA7 in ICH-induced neuronal damage. A collagenase ICH model was performed in adult male Sprague Dawley rats. First, a possible relationship between ANXA7 and ICH pathology was confirmed by an increase in the protein and mRNA level of ANXA7 in the brain tissue around hematoma of ICH rats and the rescue effects of ANXA7 knockdown in vivo on neuronal death, blood-brain barrier damage, brain edema, neurobehavioral deficient, and inflammatory response. In addition, the rescue effect of ANXA7 knockdown on neurobehavioral deficient was also verified in rat autologous blood injection ICH model. Second, we found that ICH significantly increased the phosphorylation ratio of ANXA7 at the threonine residues mainly in neurons. Finally, based on site-specific mutagenesis, we identified that ANXA7 phosphorylation at threonine 286 is required for its interaction with SNAP25 at presynaptic axon terminal and SNAP23 at postsynaptic axon terminal. Collectively, our findings suggest that ANXA7 contributed to SBI at least partially through regulating glutamate toxicity after ICH. Selective inhibition of ANXA7 phosphorylation may be a novel approach to ameliorate ICH-induced SBI.

Microfibers: Amorphous Metal‐Sulphide Microfibers Enable Photonic Synapses for Brain‐Like Computing (Advanced Optical Materials 5/2015)

The optical axon and photonic synapse transmit information with optical pulses that propagate along a fiber, whose transmission is altered through photodarkening as a result of exposure at a sub-bandgap wavelength. As explained by B. Gholipour et al. on page 635, this photomodifiation plays the role of either inhibitory or excitatory action potentials in the post-synaptic axon.

Blocking Effects of Human Tau on Squid Giant Synapse Transmission and Its Prevention by T-817 MA

Filamentous tau inclusions are hallmarks of Alzheimer's disease and related neurodegenerative tauopathies, but the molecular mechanisms involved in tau-mediated changes in neuronal function and their possible effects on synaptic transmission are unknown. We have evaluated the effects of human tau protein injected directly into the presynaptic terminal axon of the squid giant synapse, which affords functional, structural, and biochemical analysis of its action on the synaptic release process. Indeed, we have found that at physiological concentration recombinant human tau (h-tau42) becomes phosphorylated, produces a rapid synaptic transmission block, and induces the formation of clusters of aggregated synaptic vesicles in the vicinity of the active zone. Presynaptic voltage clamp recordings demonstrate that h-tau42 does not modify the presynaptic calcium current amplitude or kinetics. Analysis of synaptic noise at the post-synaptic axon following presynaptic h-tau42 microinjection revealed an initial phase of increase spontaneous transmitter release followed by a marked reduction in noise. Finally, systemic administration of T-817MA, a proposed neuro-protective agent, rescued tau-induced synaptic abnormalities. Our results show novel mechanisms of h-tau42 mediated synaptic transmission failure and identify a potential therapeutic agent to treat tau-related neurotoxicity.

Stimulation of dorsal root afferents increases the excitability of ascending sensory axons in the isolated spinal cord of mature mice

The phenomenon of windup has often been used to assess excitability increases of spinal neurons induced by repetitive stimulation of nociceptive afferents. Windup has been studied in individual spinal cord neurons and in spinal motor reflexes neither of which accurately reflect the forward transmission of nociceptive signals to the brain. In addition, most in vitro studies of spinal windup have been conducted on immature or juvenile animals and it is challenging to extrapolate these results to the adult spinal cord. In the present study, we have used an in vitro whole spinal cord preparation from functionally mature mice (up to 8 weeks old) to record windup activity in ascending axons in the mid-thoracic region evoked by electrical stimulation of a lumbar or sacral dorsal root. Windup responses were observed in axons in the ipsi- and contralateral dorsolateral funiculus (iDLF and cDLF) and in the contralateral ventrolateral funiculus (cVLF). No windup responses were evoked in postsynaptic axons of the ipsilateral dorsal columns (iDC) and no postsynaptic responses were elicited in the ipsilateral ventrolateral funiculus (iVLF) or contralateral dorsal columns (cDC). Between 40% and 45% of all axons in the DLF and cVLF that responded to a single dorsal root stimulus also showed windup. The NMDA receptor antagonist MK-801 reversibly blocked such windup responses. These results illustrate that windup can be consistently recorded from ascending pathways in the mature spinal cord in vitro but also show that windup can only be elicited in a proportion of sensory axons projecting through some, but not all, ascending spinal cord pathways.

A team of potassium channels tunes up auditory neurons

Mammals harbour a diverse array of voltage-gated K+ channels. How those channels team up in a neuron to do a job is becoming clear in the principal cells of the medial nucleus of the trapezoid body (MNTB). Ian Forsythe and his colleagues have used the electrically compact nature of MNTB neurons to measure K+ currents and combined those findings with molecular biological studies to identify, characterize and localize diverse K+ channels on MNTB neurons. A series of reports, the latest by Hardman and Forsythe being in this issue of The Journal of Physiology (Hardman & Forsythe, 2009), show that the firing of action potentials in MNTB neurons is finely controlled by a team of K+ conductances with different properties. Neurons in the MNTB are part of a neuronal circuit that assesses differences in intensity at the two ears, cues that are used to determine where in the horizontal plane sounds arise (Fig. 1A). Neurons in the lateral superior olive (LSO) compare interaural intensity by summing excitation from the ipsilateral ear with inhibition from the contralateral ear. Anyone who has participated in a conversation knows that sounds change rapidly in frequency and intensity; for interaural comparisons of intensity to be useful for localizing sounds, the time of arrival of excitation and inhibition at the LSO must be matched to within a few hundred microseconds. Figure 1 The medial nucleus of the trapezoid body (MNTB) feeds information about contralateral sounds to neurons in the lateral superior olive (LSO) that compare interaural intensity by balancing ipsilateral excitation (red) with contralateral inhibition (blue) ... How are the delays in ipsi- and contralateral inputs to the LSO matched in timing when path from the contralateral ear to the LSO is longer and includes an extra synapse? Neurons from the ipsilateral cochlear nucleus (CN) bring excitation to LSO neurons through relatively fine axons. Globular bushy cells from the contralateral CN, whose axons are particularly large and rapidly conducting, innervate MNTB neurons through calyces of Held. A calyx excites an MNTB neuron with a short and constant synaptic delay, generating large, suprathreshold EPSPs. The team of K+ channels allows MNTB neurons to relay the temporal firing pattern of globular bushy cells to contralateral LSO neurons as precisely timed, glycinergic IPSPs. Which are the members of the team of K+ channels? Voltage-gated K+ channels are formed a tetrameric combinations of α subunits from within a class of channels (e.g. Kv1). Figure 1B summarizes what is known about the presence of various α subunits in the MNTB. (1) Channels of the Kv1 family, located in the pre- and postsynaptic axons as well as on somas of MNTB neurons, provide a prominent, rapidly activating, low-voltage-activated K+ current whose activation around rest shapes synaptic potentials and prevents repetitive firing in both the calyceal terminals and in MNTB neurons (Dodson et al. 2003). (2) Channels that contain Kv2.2 are prominent in the initial segments of axons of MNTB neurons. These high-voltage-activated, slowly activating and deactivating channels hyperpolarize neurons between action potentials when they fire rapidly. In promoting repolarization between action potentials, these channels enhance Na+ currents by removing inactivation when MNTB neurons fire rapidly (Johnston et al. 2008b). (3) Channels containing Kv3.1b mediate a prominent high-voltage-activated, rapidly activating and inactivating K+ current that repolarizes action potentials, making them narrow. Kv3.1b channels are in calyceal terminals as well as on the somatic membranes of MNTB neurons. When they are phosphorylated, Kv3.1b subunits have a lower open probability. Rapid firing induced by electrical stimulation or by a noisy acoustic environment causes a decrease in phosphorylation and an increase in this K+ current (Song et al. 2005). (4) Kv4.3 subunits have been found in the MNTB of mice but not rats (Johnston et al. 2008a). Channels of the Kv4 family mediate A-currents. These K+ currents inactivate rapidly at relatively hyperpolarized potentials making them transient and most evident when depolarization is preceded by hyperpolarization. This small current would be expected to shorten and diminish EPSPs early in a train but becomes inactivated with strong stimulation. (5) Now Hardman and Forsythe show that Kv11 channels encoded by an ether-a-go-go-related gene (ERG) are also present in MNTB neurons of mice. These channels inactivate more rapidly than they activate so that they conduct current mainly upon repolarization after inactivation is removed but before they deactivate. The current through ERG channels repolarizes MNTB neurons, its action being most prominent near the resting potential after a depolarization. K+ channels work as a team to sharpen EPSPs, to sharpen action potentials, and to assure that repolarization removes Na+ inactivation even during rapid firing. K+ channels enable MNTB neurons to fire robustly and to signal with temporal precision even when they are driven at high rates.

Synapse formation between isolated axons requires presynaptic soma and redistribution of postsynaptic AChRs.

The involvement of neuronal protein synthetic machinery and extrinsic trophic factors during synapse formation is poorly understood. Here we determine the roles of these processes by reconstructing synapses between the axons severed from identified Lymnaea neurons in cell culture, either in the presence or absence of trophic factors. We demonstrate that, although synapses are maintained between isolated pre- and postsynaptic axons for several days, the presynaptic, but not the postsynaptic, cell body, however, is required for new synapse formation between soma-axon pairs. The formation of cholinergic synapses between presynaptic soma and postsynaptic axon requires gene transcription and protein synthesis solely in the presynaptic neuron. We show that this synaptogenesis is contingent on extrinsic trophic factors present in brain conditioned medium (CM). The CM-induced excitatory synapse formation is mediated through receptor tyrosine kinases. We further demonstrate that, although the postsynaptic axon does not require new protein synthesis for synapse formation, its contact with the presynaptic cell in CM, but not in defined medium (no trophic factors), differentially alters its responsiveness to exogenously applied acetylcholine at synaptic compared with extrasynaptic sites. Together, these data suggest a synergetic action of cell-cell signaling and trophic factors to bring about specific changes in both pre- and postsynaptic neurons during synapse formation.

Neuronal nicotinic acetylcholine receptor in the hypothalamus: morphological diversity and neuroendocrine regulations

The subcellular localization and functional significance of neuronal nicotinic acetylcholine receptor α 4-subunits were investigated in the rat hypothalamic supraoptic nucleus. A high level of α 4 mRNA expression was found in the magnocellular neurons in the supraoptic nucleus. Strong immunoreactitivy for α 4 in neurons of the supraoptic nucleus was detected in the rough endoplasmic reticulum and cytoplasmic matrix, although it was very weak in the Golgi apparatus, except for the transport vesicles. Immunoreactivity for α 4 was detected in both the pre-synaptic axon terminals and post-synaptic axon terminals. A high level of signals for vasopressin mRNA was detected in the supraoptic nucleus after the animals were injected s.c. with nicotine. These findings suggest that α 4-containing subtypes are synthesized in the rough endoplasmic reticulum and transported to the plasma membrane and serve as pre- and post-synaptic nicotinic acetylcholine receptors. Nicotine may up-regulate vasopressin gene expression in the supraoptic nucleus, acting through nicotinic acetylcholine receptors.

Localization, physiology, and modulation of a molluskan dopaminergic synapse

We investigated the location, physiology, and modulation of an identified synapse from the central nervous system (CNS) of the mollusk Lymnaea stagnalis. Specifically, the excitatory synapse from interneuron right pedal dorsal one (RPeD1) to neurons visceral dorsal two and three (VD2/3) was examined. The gross and fine morphology of these neurons was determined by staining with Lucifer yellow or sulforhodamine. In preparations where RPeD1 was stained with Lucifer yellow and VD2/3 with sulfo-rhodamine, the axon collaterals occupied similar regions, suggesting that these neurons make physical contact in the CNS. Digital confocal microscopy of these preparations revealed that presynaptic varicosities made apparent contact (synapses) with smooth postsynaptic axon collaterals. The number of putative synapses per preparation was about five to 10. Regarding physiology, the synaptic latency was moderately rapid at 24.1 ± 5.2 ms. Previous work indicated that RPeD1 uses dopamine as a neurotransmitter. The RPeD1 → VD2/3 excitatory postsynaptic potential (EPSP) and the VD2/3 bath-applied dopamine (100-μM) response displayed a similar decrease in input resistance and a similar predicted reversal potential (−31 vs. −26 mV), indicating that the synapse and exogenous dopamine activate the same conductance. Finally, bath-applied serotonin (10 μM) rapidly and reversibly depressed the RPeD1 → VD2/3 synapse but did not affect the VD2/3 bath-applied dopamine (100-μM) response, suggesting a presynaptic locus of action for serotonin. The effect of serotonin was not associated with any changes to the pre- or postsynaptic membrane potential and input resistance, or the presynaptic action potential half-width. The RPeD1 → b3 VD2/3 synapse provides an opportunity to examine the anatomy and physiology of transmission, and is amenable to the study of neuromodulation. © 1997 John Wiley & Sons, Inc. J Neurobiol 33: 247–264, 1997

Localization, physiology, and modulation of a molluskan dopaminergic synapse

We investigated the location, physiology, and modulation of an identified synapse from the central nervous system (CNS) of the mollusk Lymnaea stagnalis. Specifically, the excitatory synapse from interneuron right pedal dorsal one (RPeD1) to neurons visceral dorsal two and three (VD2/3) was examined. The gross and fine morphology of these neurons was determined by staining with Lucifer yellow or sulforhodamine. In preparations where RPeD1 was stained with Lucifer yellow and VD2/3 with sulforhodamine, the axon collaterals occupied similar regions, suggesting that these neurons make physical contact in the CNS. Digital confocal microscopy of these preparations revealed that presynaptic varicosities made apparent contact (synapses) with smooth postsynaptic axon collaterals. The number of putative synapses per preparation was about five to 10. Regarding physiology, the synaptic latency was moderately rapid at 24.1 +/- 5.2 ms. Previous work indicated that RPeD1 uses dopamine as a neurotransmitter. The RPeD1 --> VD2/3 excitatory postsynaptic potential (EPSP) and the VD2/3 bath-applied dopamine (100-microM) response displayed a similar decrease in input resistance and a similar predicted reversal potential (-31 vs. -26 mV), indicating that the synapse and exogenous dopamine activate the same conductance. Finally, bath-applied serotonin (10 microM) rapidly and reversibly depressed the RPeD1 --> VD2/3 synapse but did not affect the VD2/3 bath-applied dopamine (100-microM) response, suggesting a presynaptic locus of action for serotonin. The effect of serotonin was not associated with any changes to the pre- or postsynaptic membrane potential and input resistance, or the presynaptic action potential half-width. The RPeD1 --> VD2/3 synapse provides an opportunity to examine the anatomy and physiology of transmission, and is amenable to the study of neuromodulation.

The inositol high-polyphosphate series blocks synaptic transmission by preventing vesicular fusion: a squid giant synapse study.

Presynaptic injection of inositol 1,3,4,5-tetraphosphate, inositol 1,3,4,5,6-pentakisphosphate, or inositol 1,2,3,4,5,6-hexakisphosphate--which we denote here the inositol high-polyphosphate series (IHPS)--is shown to block synaptic transmission when injected into the preterminal of the squid giant synapse. This effect is not produced by injection of inositol 1,4,5-trisphosphate. The synaptic block is characterized by a time course in the order of 15-45 min, depending on the injection site in the preterminal fiber; the fastest block occurs when the injection is made at the terminal release site. Presynaptic voltage clamp during transmitter release demonstrates that IHPS block did not modify the presynaptic inward, calcium current. Analysis of synaptic noise at the postsynaptic axon shows that both the evoked and spontaneous transmitter release are blocked by the IHPS. Tetanic stimulation of the presynaptic fiber at frequencies of 100 Hz indicates that block is accompanied by gradual reduction of the postsynaptic response, demonstrating that the block interferes with vesicular fusion rather than with vesicular docking. These results, in combination with the recently demonstrated observation that the IHPS bind the C2B domain in synaptotagmin [Fukada, M., Aruga, J., Niinobe, M., Aimoto, S. & Mikoshiba, K. (1994) J. Biol. Chem. 269, 29206-29211], suggest that IHPS elements are involved in vesicle fusion and exocytosis. In addition, a scheme is proposed in which synaptotagmin triggers transmitter release directly by promoting the fusion of synaptic vesicles with the presynaptic plasmalemma, in agreement with the very rapid nature of transmitter release in chemical synapses.

Light- and electron-microscopic analysis of neuropeptide Y-immunoreactive profiles in the cat spinal dorsal horn

The organization of neuropeptide Y-containing profiles in the dorsal horn of cat lumbosacral spinal cord was examined in an immunocytochemical study employing a specific antiserum against neuropeptide Y. Light-microscopic inspection revealed heavy concentrations of immunoreactive axons and varicosities within the superficial layers of the dorsal horn (laminae I and II) and only low to moderate numbers of positive terminals in the deeper layers (laminae III–VI). Neuropeptide-Y immunoreactivity in the superficial laminae occurred primarily as single punctate terminals, although in sagittal sections long rostrocaudally orientated fibres were also found. Immunoreactive fibres in the deeper layers were usually long and beaded. Two-hundred and eight neuropeptide Y-immunoreactive profiles throughout laminae I–VI were examined through serial sections with the electron microscope, and the overwhelming majority ( n = 194) was confirmed to be axon terminals, most of which (95%) formed synaptic junctions. These terminals were packed with small irregularly shaped agranular vesicles, together with a number of large dense-core vesicles. Immunoreactivity was homogenously scattered throughout the cytoplasm, and was also associated with the dense-core vesicles. A few neuropeptide Y-containing profiles ( n = 14) were difficult to classify but they could have been vesicle-containing dendrites. The postsynaptic targets of neuropeptide Y-positive terminals were similar throughout each dorsal horn lamina. Most frequently, neuropeptide Y-positive boutons formed axodendritic and axosomatic synaptic junctions (range = 64% of synapses in laminae V/VI to 83% in lamina III). A smaller proportion of synapses were found upon other axon terminals and in laminae I–III the postsynaptic axon terminals were sometimes the central boutons of glomeruli. A number of terminals, especially those in lamina II, formed multiple synapses which often comprised a triadic arrangement. These findings suggest that neuropeptide Y regulates spinal sensory transmission through both a postsynaptic action upon dorsal horn neurons and a presynaptic action upon primary afferent terminals.

Possible morphological substrates for GABA-mediated presynaptic inhibition in the lamprey spinal cord.

Gamma-aminobutyric acid (GABA) neurons intrinsic to the lamprey spinal cord are known to modulate synaptic transmission from interneurons active during locomotion and from mechanosensory dorsal cells. Many of these physiological effects are presynaptic. To establish the morphological substrates for these axo-axonic interactions, an ultrastructural analysis was performed with an antiserum to fixed GABA. The GABA immunoreactivity (ir) was detected by postembedding peroxidase-antiperoxidase and immunogold techniques. GABA-ir terminals were found to make appositions with unlabelled axons located in the dorsal columns and in the ventrolateral aspect of the spinal cord. In the ventrolateral part of the cord, similar appositions between different GABA-ir terminals were also observed. The immunolabelled terminals contained spherical to pleomorphic synaptic vesicles, and also glycogen granules and dense core vesicles. In some cases, the fine structure of the contacts between immunogold-labelled terminals and unlabelled axons suggested a synaptic relationship. Such a relation was found in a relatively small proportion (2-3%) of the appositions studied. These specializations were always observed in close relation to an output synapse of the postsynaptic axon. It is suggested that the axo-axonal contacts described may provide an effective modulation of the synaptic transmission from axons in the lamprey spinal cord.

Neuropeptide Y-immunoreactive terminals form axo-axonic synaptic arrangements in the substantia gelatinosa (lamina II) of the cat spinal dorsal horn

The ultrastructural organization of nerve terminals containing neuropeptide Y-immunoreactivity was studied in the substantia gelatinosa of the cat spinal dorsal horn. Seventy immunoreactive boutons were examined through serial sections and 67 of them were found to form between one and five synaptic junctions with dendrites (59.5% of synapses), somata (3% of synapses) and other axon terminals (37.5% of synapses). The postsynaptic axon terminals were often the central boutons of glomeruli. These findings suggest that neuropeptide Y regulates spinal sensory transmission through both a postsynaptic action upon dorsal horn neurons and a presynaptic action upon primary afferent terminals.

Electrically evoked turning: asymmetric and symmetric collision between anteromedial cortex and striatum

Electrical stimulation of the anteromedial cortex (AMC) or striatum of rats evoked contraversive eye, head and body movements. In these experiments we test which neurons and which pathways are responsible for the turning by delivering conditioning (C) pulses to one site and test (T) pulses to the second site, and measuring the frequency of pulse pairs required to evoke a full turn in 10 s. Decreases in the required frequency were usually found at C-T intervals from 0.6 to 1.0 ms, whether the C pulses were delivered to the AMC or to the striatum. This symmetric effect is attributed to collision in fast-conducting axons connecting cortex and striatum. Symmetric collision at C-T intervals of 2–4 ms was observed between cortex and 3 dorsal striatal sites, suggesting slower axons from cortex to these dorsal striatal sites. In several animals, asymmetric changes in required frequency also occurred. When the C pulses were presented via the striatal electrode, the recovery in required frequency occurred at C-T intervals of 1–4 ms, but when the C pulses were presented via the cortical electrode, recovery occurred at C-T intervals of 2–50 ms. This asymmetry is attributed to indirect (i.e., transynaptic) activation of corticostriatal or striatal output axons. These results suggest that in both cortex and striatum there are synapses, transmitting from rostral to caudal, which are important for electrically evoked turning. When C and T pulses were delivered to the same site, decreases in required frequency occurred at C-T intervals from 0.4 to 4 ms, attributable to recovery from refractoriness. In 3 striatal sites, however, large changes were also seen at C-T intervals from 6 to 50 ms. In all 3 sites, asymmetric collision occurred at these same intervals. The recovery at long C-T intervals could be due to transynaptic collision also, resulting from the simultaneous activation of presynaptic and postsynaptic axons by a single striatal electrode.

Computer study of presynaptic inhibition controlling the spread of action potentials into axonal terminals

1. The effect of presynaptic, axoaxonal inhibition, that exerts its action by producing a local conductance increase, on the behavior of action potentials at postsynaptic axon terminals is analyzed computationally. The significance of the location and strength of the presynaptic inhibition, as well as the morphology and membrane properties of the axonal terminals, are considered. 2. Keeping the specific properties of terminal membrane and axoplasm constant, the critical "silent" steady-state conductance change (gcrit) that blocks propagation is linearly scaled with the terminal diameter raised to the 3/2 power. At the midpoint of a 5 lambda long, 1 micron diameter axon that has the standard Hodgkin and Huxley (1952) kinetics at 18 degrees C (and an input conductance of 8.7 nS), gcrit is 72 nS. At 0 degrees C, gcrit = 200 nS, whereas at 30 degrees C gcrit = 30 nS. 3. The critical conductance change that blocks propagation depends steeply on the density of excitable channels (gNa) at the terminal. For a geometrically uniform terminal at 18 degrees C, the action potential can not be blocked by a local shunt when gNa greater than 600 mS/cm2. 4. An axoaxonal synapse in the proximity of the postsynaptic release site has graded control over the spike amplitude (and, therefore, over the postsynaptic output) at that site. Presynaptic inhibition located remotely from the release site has an all-or-none effect at the release site. 5. Inhibition is more effective in attenuating the spike at the terminal when it impinges onto a passive terminal rather than on an excitable one. 6. The detectability of the conductance increase as well as the change in action potential amplitude associated with the presynaptic inhibition is poor at only a short distance from the axoaxonal synapse. The presence of bottlenecks and varicosities at some axonal terminals enhances this decoupling effect even more. Thus presynaptic inhibition may reduce the potential near the output site of the affected axon significantly, without producing any noticeable conductance or voltage change at a distance of only several tens of micrometers (a few tenths of a space constant) away from it. 7. Varicosities and bottlenecks typically found along axonal terminals are the optimal loci for presynaptic inhibition to be most effective in attenuating the action potential at the terminal. Even with high density of excitable channels, propagation with such terminal geometry is insecure, and one, or few, quanta of inhibitory transmitter (a conductance change of several nanosiemens) can block propagation there.(ABSTRACT TRUNCATED AT 400 WORDS)

The actions of L-glutamate at the postsynaptic membrane of the squid giant synapse.

The actions of L-glutamate on the postsynaptic membrane of the squid giant synapse were investigated using two methods of application: ionophoresis and bath perfusion. Bath perfusion of 10 mmoll-1 sodium glutamate did not produce an appreciable depolarization of the postsynaptic membrane but reversibly blocked the neurally evoked postsynaptic potential (PSP). The postsynaptic membrane depolarized when L-glutamate was applied ionophoretically. The sensitivity to glutamate application was not uniform, but sharply localized to sites which may correspond to synaptic contacts made by branching colaterals from the postsynaptic axon. The relationship between membrane potential and amplitude of the glutamate-activated postsynaptic potential (PSP) examined under current-clamp conditions was linear over the voltage range studied (-110 to -60 mV) with an extrapolated reversal potential of -36 mV. The amplitude of the glutamate-activated PSP was reduced either by replacing Na+ in the external solution with Tris+ (Na+-free) or by raising the extracellular K+ concentration to 20 mmoll-1 and was abolished by removing both Na+ and Ca2+ from the bath solution. The PSP amplitude was insensitive to changes in the extracellular Mg2+ concentration. The extrapolated reversal potential of the glutamate PSP was shifted to more positive potentials in both Na+-free and raised-K+ bathing solutions and was unchanged by anion substitution. The depolarization induced by L-glutamate increased with increasing ionophoretic current and reached a maximum with large pulses. Double logarithmic plots of the coulomb dose-response relationship gave a limiting slope in the range 1.7-2.2, suggesting that two glutamate molecules are required for receptor activation. The time course of desensitization of the glutamate response was studied using a double-pulse method. The initial decrease in the ratio, PSP2/PSP1, is followed by a slower time-dependent recovery of the postsynaptic response with a time constant of 8.5 s. Prolonged perfusion of the squid giant synapse with concanavalin A failed to abolish desensitization of the glutamate-evoked PSP.

The form and dimensions of the giant synapse of squids

A study was made of the distal giant synapse, and of proximal synapses, in the stellate ganglion of the squid, Loligo vulgaris . For this purpose we injected iontophoretically dyes or cobalt ions into the pre- or postsynaptic axon. The intra-axonal movement of visible dyes was measured. Both presynaptic fibres, the main second order giant axon and the largest accessory axon, branched to make multiple synaptic contacts on the giant motor axons from near the perikarya down to near the exit of the stellar nerves from the ganglion. There were considerable individual variations in the branching patterns of the presynaptic giant axon and in the course and number of the postsynaptic giant axons. More than one accessory axon made contact with the largest motor axon. Fine structural details of the synapse are presented. The size of the contact area made by the main presynaptic axon on the last postsynaptic axon of a medium-sized animal was estimated from low power electron micrographs. We measured and counted synaptic contacts, synaptic vesicles and mitochondria. The fine structure of proximal synapses was found to be very similar to that of the distal synapse. Cobalt- or dye-injected ganglia showed that the perikarya of the axons which fuse to form the postsynaptic giant axons are located in diffuse and overlapping areas of the giant fibre lobe. In freshly hatched larvae the giant synapse was well differentiated; a gradient of differentiation from brain to periphery was detectable. The distal giant synapses of the oegopsid squid Todarodes sagittatus and of Sepia officinalis differed from the Loligo synapse. In Todarodes and Sepia collaterals and processes from both the presynaptic and the postsynaptic giant fibres are shown to meet in numerous contacts in the enlarged sheath surrounding the third order axon. In several respects the Loligo giant fibre system appears to represent in phylogenetical order the more evolved neuronal network.

Post-synaptic calcium influx at the giant synapse of the squid during activation by glutamate.

Changes in free calcium were monitored in the post-synaptic axon of the giant synapse of the squid, using the calcium indicators aequorin and Arsenazo III. The peak size of the calcium-dependent optical signals recorded from aequorin and Arsenazo III both showed a linear relation with the amount of calcium injected ionophoretically into the axon, but the Arsenazo signal had a slower time course than the aequorin. Ionophoretic application of glutamate to the post-synaptic axon depolarized the axon and caused a rise in intracellular free calcium. Aequorin signals were detected in natural sea water, and their size increased when the calcium concentration in the sea water was raised. Arsenazo signals could be detected only in high-calcium (55 mM) sea water. Intracellular calcium signals were detected also during bath application of several glutamate analogues, including kainate, ibotenate, and aspartate. The peak amplitude of the intracellular calcium signal, monitored with both indicators, increased with increasing ionophoretic glutamate dose, and varied linearly with the integral of the glutamate-induced membrane depolarization. No calcium signals were detected when depolarizations, similar to those produced by glutamate, were induced by current injection in the absence of glutamate. We conclude that glutamate increases the calcium permeability of the post-synaptic membrane, independently of the glutamate-induced depolarization. The glutamate-induced depolarization and the rise in intracellular free calcium increased roughly linearly as the membrane potential was made more negative. Extrapolation of these data indicated that the glutamate depolarization would reduce to zero at about -30 mV, while the calcium signals would be suppressed at about +50 mV.

Receptive field organization and response properties of spinal neurones with axons ascending the dorsal columns in the cat.

Micro-electrode recordings were made from single post-synaptic axons in the dorsal columns of cats anaesthetized with chloralose and paralysed with gallamine triethiodide. The recordings were made from the L5 segment and the axons were shown to project to the upper cervical level. Forty-eight units were recorded and the axons had conduction velocities of 22-61 ms-1, averaging 38.3 ms-1. Excitatory receptive fields were complex in many units, being made up of clearly defined, separate, low and high threshold areas. The receptive fields were often discontinuous. Only a few units behaved as if they received excitatory input from a single class of mechanoreceptors. A minority (13%) of units had labile, excitatory receptive fields that expanded in size during the recording period. About 40% of the units had inhibitory receptive fields. These were of two main types: either small and within or adjacent to the excitatory field, or large and separated from or adjacent to the excitatory field. The great majority of units had resting discharges upon isolation and these consisted of single impulses or bursts of impulses at short intervals separated by longer, irregular periods. The time course of inhibition produced by electrical stimulation of cutaneous nerves suggested presynaptic inhibitory components to the inhibition. Some inhibitory curves were very prolonged with maxima at about 100 ms and total durations of up to 400 ms. The complexity of the receptive field organization in these dorsal horn neurones is discussed, as is their possible significance as input neurones to the dorsal column nuclei.