Axon Hillock Research Articles

Impairment of axonal transport in the axon hillock and the initial segment of anterior horn neurons in transgenic mice with a G93A mutant SOD1 gene

Impaired axonal transport of the fast or slow component has been reported in patients with sporadic amyotrophic lateral sclerosis (ALS), animal models for ALS, and familial ALS-linked mutant Cu/Zn superoxide dismutase (SOD1) transgenic mice. However, little is known about the impairment of axonal transport in mutant SOD1 transgenic mice. This is the first electron microscopic investigation of the axon hillock (AH) and the initial segment (IS) of anterior horn cells in the spinal cord of transgenic mice expressing the G93A mutant human SOD1, and it was launched with a view toward examining whether the axonal transport is impaired in this region. Six transgenic mice were killed at ages ranging from the presymptomatic to symptomatic stages. Six age-matched non-transgenic wild-type mice served as controls. In the non-transgenic mice, 91 AH and IS were observed, but those with increased neurofilaments or mitochondria were rarely found. In the transgenic mice, 95 AH and IS directly emanating from normal-looking large anterior horn cells were seen. AH and IS with increased neurofilaments or, to a lesser extent, increased mitochondria, and round-shaped mitochondria in particular, were more frequently observed, even at the early presymptomatic stage, than in the controls, and the frequency increased with time through the presymptomatic stages. On the other hand, the somata of large motor neurons directly connected with the axons did not exhibit any abnormal accumulation of neurofilaments or mitochondria. These findings suggest that both the slow axonal transport of neurofilaments and the fast axonal transport of mitochondria are impaired in AH and IS before the onset of disease in this animal model.

Spider Peripheral Mechanosensory Neurons Are Directly Innervated and Modulated by Octopaminergic Efferents

Octopamine is a chemical relative of noradrenaline providing analogous neurohumoral control of diverse invertebrate physiological processes. There is also evidence for direct octopaminergic innervation of some insect peripheral tissues. Here, we show that spider peripheral mechanoreceptors are innervated by octopamine-containing efferents. The mechanosensory neurons have octopamine receptors colocalized with synapsin labeling in the efferent fibers. In addition, octopamine enhances the electrical response of the sensory neurons to mechanical stimulation. Spider peripheral mechanosensilla receive extensive efferent innervation. Many efferent fibers in the legs of Cupiennius salei are GABAergic, providing inhibitory control of sensory neurons, but there is also evidence for other neurotransmitters. We used antibody labeling to show that some efferents contain octopamine and that octopamine receptors are concentrated on the axon hillocks and proximal soma regions of all mechanosensory neurons in the spider leg. Synaptic vesicles in efferent neurons were concentrated in similar areas. Octopamine, or its precursor tyramine, increased responses of mechanically stimulated filiform (trichobothria) leg hairs. This effect was blocked by the octopamine antagonist phentolamine. The octopamine-induced modulation was mimicked by 8-Br-cAMP, a cAMP analog, and blocked by Rp-cAMPS, a protein kinase A inhibitor, indicating that spider octopamine receptors activate adenylate cyclase and increase cAMP concentration. Frequency response analysis showed that octopamine increased the sensitivity of the trichobothria neurons over a broad frequency range. Thus, the major effect of octopamine is to increase its overall sensitivity to wind-borne signals from sources such as flying insect prey or predators.

The role of the cytoskeleton in cell body enlargement, increased nuclear eccentricity and chromatolysis in axotomized spinal motor neurons

BackgroundWhen spinal motor axons are injured, the nucleolus, nucleus and cell body of the injured cell transiently increase in size, the nucleus becomes more eccentrically placed, and the organization of polyribosomes into Nissl bodies is temporarily disrupted. The mechanisms for these classical morphological responses to axotomy have not been satisfactorily explained.ResultsIn this study we address the role of the cell body cytoskeleton in these structural changes. We show that the cytoskeleton of uninjured lumbar motor neuron cell bodies maintains nucleolar, nuclear and cell body size and nuclear position. When isolated, the relatively insoluble cell body cytoskeleton contains Nissl bodies and lipofuscin granules. After axotomy, protein labeling increases markedly and the cytoskeleton enlarges, increasing nucleolar, nuclear and cell body size, as well as nuclear eccentricity. Nearly all of the protein mass that accumulates in the cell body after axotomy appears to be added to the cytoskeleton.ConclusionWe conclude that axotomy causes the conjugate enlargement of the nucleolus, nucleus and cell body and increases nuclear eccentricity in spinal motor neurons by adding protein to the cytoskeleton. The change in nuclear position, we propose, occurs when cytoskeletal elements of the axon cannot enter the shortened axon and "dam up" between the nucleus and axon hillock. As a consequence, we suggest that Nissl body-free axonal cytoskeleton accumulates between the nucleus and axon, displaces Nissl body-containing cytoskeleton, and produces central chromatolysis in that region of the cell.

Neuronal gap junctions in the mouse main olfactory bulb: Morphological analyses on transgenic mice

In the present study we analyzed the structural features of extraglomerular gap junction-forming processes in mouse olfactory bulb electron microscopically. This work complements a previous study in which we analyzed the structural features of neuronal gap junction-forming processes within the glomerulus itself. Furthermore we examined connexin 36 expressing cells in the mouse olfactory bulb by analyzing transgenic mice in which the connexin 36 coding sequence was replaced with histological reporters. In extraglomerular regions, the mitral/tufted cell somata, dendrites and axon hillocks made gap junctions and mixed synapses with interneuronal processes. These gap junctions and synapses were associated with various types of interneuronal processes, including a particular type of sheet-like or calyx-like process contacting the somata or large dendrites of mitral/tufted cells. In the olfactory bulbs of the transgenic mice, connexin 36 was expressed in mitral cells, tufted cells, presumed granule cells and periglomerular cells. Multiple immunofluorescent labelings further revealed that presumed interneurons expressing connexin 36 in the periglomerular region rarely expressed calbindin, calretinin or tyrosine hydroxylase and are likely to comprise a chemically uncharacterized class of neurons. Similarly, interneurons expressing connexin 36 in the granule cell layer were rarely positive for calretinin, which was expressed in numerous presumed granule cells in the mouse main olfactory bulb. In summary, these findings revealed that mitral/tufted cells make gap junctions with diverse types of neurons; in the glomeruli gap junction-forming interneuronal processes originated from some types of periglomerular cells but others from a hitherto uncharacterized neuron type(s), and in the extraglomerular region gap-junction forming processes originate mainly from a subset of cells within the granule cell layer.

Differential sorting and packaging of capa-gene related products in an insect

A unique costorage of neuropeptides was recently found in the abdominal perisympathetic organs (PSOs) of the American cockroach, Periplaneta americana. Having specific antisera directed against all peptides belonging to this neurosecretory system, we examined the sorting of PSO-peptides in the soma of the median neurosecretory cells of abdominal ganglia by using immunoelectron microscopic double stainings. The data indicate that all six abundant neuropeptides of this neurohormonal system, which includes three capa-gene related products, are primarily incorporated into separate vesicles. These vesicles fuse with each other in the cytoplasm and become translucent on their way to the axon hillock. By means of light microscopy and MALDI-TOF mass spectrometry, an identical population of neuropeptides was found in interneurons of the brain. As revealed by subsequent immunoelectron microscopic analysis, the peptides of these cells are separately packed into dense core vesicles but do not fuse with each other. Thus, hitherto unknown cell-type-specific sorting mechanisms occur in neurosecretory cells and interneurons, respectively.

Ganglioside GD1a increases the excitability of voltage-dependent sodium channels

The effect of the negatively charged ganglioside GD1a, one of the major brain gangliosides [H. Beitinger, W. Probst, R. Hilbig, H. Rahmann, Seasonal variability of sialo-glycoconjugates in the brain of the Djungarian hamster ( Phodopus sungorus). Comp. Biochem. Physiol., B 86 (1987) 377-384] on the function of brain derived BTX-modified voltage-dependent sodium channel was studied using the planar lipid bilayer system. Bilayers were formed either with a mixture of neutral phospholipids (4 phosphoethanolamine (PE):1 phosphocholine (PC)) alone or with one containing 6% of the disialoganglioside GD1a. The permeation and activation properties of the channels were measured in the presence of symmetrical 200 mM NaCl. We found that the single channel conductance was not affected by GD1a, whereas the steady-state activation curve displayed a hyperpolarizing shift in the presence of GD1a. Since the lipid distribution in these membranes is symmetrical, then the GD1a effect on sodium channels may result either from an induction of channel conformational changes or from an asymmetrical interaction between the channel (extracellular vs. intracellular channel aspect) and GD1a. Regardless of the mechanism, the data indicate that differences in ganglioside content in neuronal cells may contribute to the previously observed sodium channel functional variability within (soma, dentritic, axon hillock) and between neuronal cells as well as to excitability changes in those physiological and pathological conditions where changes in the neuronal ganglioside content occur.

Evidence for an axonal localization of the type 2 corticotropin-releasing factor receptor during postnatal development of the mouse cerebellum

Previous studies have described the embryonic and postnatal development of CRF, as well as the type 1 CRF receptor in the mouse cerebellum. The present immunohistochemical study localizes the cellular distribution of the type 2 CRF receptor (CRF-R2) during postnatal development of the mouse cerebellum. Western blot analysis indicates that the antibody used in this analysis recognizes both a full-length and a truncated isoform of the type 2 receptor. We propose that each isoform has a unique cellular distribution. In the present study, the postnatal (P) development (P0–P14) and cellular localization of CRF-R2 in different cell types was analyzed using PAP and double-label fluorescent immunohistochemistry; cell-specific antibodies were used to identify cells expressing CRF-R2 at different stages of postnatal development. At P0, CRF-R2 immunoreactivity was localized within the somata of Purkinje cells and migrating GABAergic interneurons. CRF-R2 was first observed in the initial axonal segments of some Purkinje cells at P5, and was evident in many Purkinje cell axon hillocks at P8. Punctate immunoreactivity is present in the molecular layer by P5 and is interpreted to be immunolabeled parallel fibers. Between P8 and P14, CRF-R2 immunostaining is present in the initial axonal segments of Golgi cells, within the internal granule cell layer. Finally, CRF-R2 is present in both radial glia in the molecular layer as well as in astrocytes in the white matter and internal granule cell layer from P5 to P14. The present results suggest that CRF-R2, both the truncated and the full-length isoforms, are present in the developing cerebellum, each with a unique cellular distribution. The immunohistochemical evidence indicates that the truncated isoform of the type 2 CRF receptor is in the axons of several different types of cerebellar cortical neurons, and suggests that CRF could play a role in cerebellar development by modulating the release of transmitters from excitatory and/or inhibitory interneurons, which in turn could directly alter the maturation of cerebellar circuits. In contrast, the binding of a ligand to the full-length isoform of CRF-R2 or to CRF-R1, both in a postsynaptic location, may have a more direct effect on regulating the responsiveness of these cells to growth factors or neurotransmitters released from afferent axons by regulating permeability of ion channels or altering second messenger systems.

Selective synaptic cadherin expression by traced neurons of the chicken visual system

The stable and specific locking-in of pre- and postsynaptic membranes in synaptogenesis may be mediated by integral membrane proteins, such as members of the cadherin family. Cadherins are ideal candidate molecules for mediating synaptic specificity because they are differentially expressed in functionally connected brain structures. We studied the expression of four classic cadherins (R-cadherin, N-cadherin, cadherin-6B and cadherin-7) at the synaptic level on the somata and the proximal neurites of identified neuron populations that were traced selectively in the developing chicken visual system. Three major findings were observed. (1) Synapses on somata of shepherd's crook cells of the optic tectum are associated preferentially with one cadherin subtype. (2) In an isthmic nucleus that contains a mixed population of cells expressing different cadherins, somatic synapses tend to express the same cadherin subtype as the rest of the cell. (3) In the oculomotor complex, two cadherin subtypes are expressed only by synapses on the axon hillock. However, another neuron type that projects from the tectum to the isthmic nucleus does not show such selective synaptic cadherin staining. Our findings support the idea that a cadherin-based adhesive mechanism can mediate synaptic specificity.

Control serotoninérgico de la corteza prefrontal

The prefrontal cortex (PFC) plays a crucial role in higher brain functions such as working memory or cognition and controls, via the excitatory axons of pyramidal neurons, the activity of many subcortical motor and limbic areas. It receives a dense innervation from the brainstem aminergic nuclei, including the serotonergic raphe nuclei. Prefrontal function and metabolism is altered in patients with severe psychiatric disorders, like major depression or schizophrenia. Although the exact role of serotonergic neurotransmission in PFC remains largely unknown, the PFC contains a very large density or serotonin 5-HT1A (inhibitory) and 5-HT2A (excitatory) receptors. In addition, hallucinogens like LSD or DOI are agonists and atypical antipsychotics are antagonists at 5-HT2A receptors. In this review we focus on the main excitatory and inhibitory mechanisms through which serotonin modulates pyramidal and GABAergic neuron activity in the PFC. We report on the presence of 5-HT1A and 5-HT2A receptor-mediated responses in pyramidal neurons of the PFC that exert opposite effects on their activity when recorded in vivo in the anesthetized rat. Despite the large co-expression of both receptors in pyramidal neurons of the PFC, physiological amounts of 5-HT mainly inhibit pyramidal neurons. This is probably due to the distinct location of 5-HT1A and 5-HT2A in pyramidal neurons. Thus, 5-HT1A receptors are mainly localized in the axon hillock, where they may have a prominent inhibitory role in the control of pyramidal activity given their coupling to GIRK channels. Moreover, 5-HT can inhibit pyramidal neurons indirectly through the activation of 5-HT2A and 5-HT3 receptors localized in GABAergic interneurons and a subsequent increase in synaptic GABA inputs.

Peripheral Hot Spots for Local Ca 2+ Release after Single Action Potentials in Sympathetic Ganglion Neurons

Ca 2+ release from the endoplasmic reticulum (ER) contributes to Ca 2+ transients in frog sympathetic ganglion neurons. Here we use video-rate confocal fluo-4 fluorescence imaging to show that single action potentials reproducibly trigger rapidly rising Ca 2+ transients at 1–3 local hot spots within the peripheral ER-rich layer in intact neurons in fresh ganglia and in the majority (74%) of cultured neurons. Hot spots were located near the nucleus or the axon hillock region. Other regions exhibited either slower and smaller signals or no response. Ca 2+ signals spread into the cell at constant velocity across the ER in nonnuclear regions, indicating active propagation, but spread with a ( time) 1/2 dependence within the nucleus, consistent with diffusion. 26% of cultured cells exhibited uniform Ca 2+ signals around the periphery, but hot spots were produced by loading the cytosol with EGTA or by bathing such cells in low-Ca 2+ Ringer's solution. Peripheral hot spots for Ca 2+ release within the perinuclear and axon hillock regions provide a mechanism for preferential initiation of nuclear and axonal Ca 2+ signals by single action potentials in sympathetic ganglion neurons.

Immunocytochemical localization of reelin in the olfactory bulb of the heterozygous reeler mouse: An animal model for schizophrenia

Because heterozygous reeler (HR) mice share some abnormal traits with schizophrenic patients, and schizophrenia is often accompanied by impairment of olfactory function, this study examines reelin in the olfactory bulb of the HR mouse. In the WT mouse, reelin immunoreactivity is found in the extracellular matrix, and in the cytoplasm of olfactory nerve fibers, GABAergic interneurons, and glutamatergic mitral cells. Western blot analysis reveals that reelin immunoreactivity in the HR mouse is reduced by 45% compared to WT mouse. This is especially evident in the glomerular GABAergic interneurons. In WT mitral cells, reelin is found in discrete clumps near the axon hillock and within the axon. In the HR mouse, reelin axonal staining is diffuse and densely packed. In the rostral migratory stream of the HR mouse, immunolabeling shows an accumulation of reelin-containing neuronal precursors, apparently unable to shift from tangential to radial migration. These observations indicate that there is a downregulation of reelin in the HR mouse and suggest that secretion of reelin may be compromised. Further studies of the HR mouse may provide a new basis for understanding the role of reelin in the adult CNS, especially as it may relate to schizophrenia.

Synaptology of the rostral reticular thalamic nucleus of absence epileptic WAG/Rij rats

The adult WAG/Rij rat is a well-established animal model for human absence epilepsy characterized by the presence of spike-wave discharges (SWDs). The pacemaking activity of the rostral reticular thalamic nucleus (rRTN) has been demonstrated to be essential for SWD maintenance. We investigated if SWD maintenance can be related to the synaptic organization of the rRTN, by studying the ultrastructure of the rRTN of absence epileptic WAG/Rij rats in comparison with that of non-epileptic, age-matched ACI control rats. In WAG/Rij rats, D-, L- and F-type terminals constitute the synaptic organization of the rRTN. D-type synapses, especially axo-dendritic ones, occur frequently. L- and F-type terminals are common but less frequent than D-type terminals. Semi-quantitative observations indicate that all terminal types are present on different parts of the postsynaptic neuron, but in different numbers: they are frequent on dendrites, common on somata and axons, and occur occasionally on dendritic spines. In addition, occasionally an F-type terminal was observed on the axon hillock. The three terminal types are also involved in multiple synaptic configurations, convergent as well as divergent, with dendrites, somata, axon hillocks and axons as postsynaptic structures. Convergent synaptic configurations outnumber divergent ones. The synaptic organization of the rRTN of the non-epileptic ACI rat appears to be very similar to that of the epileptic WAG/Rij rat. This indicates that SWD maintenance in the WAG/Rij rat does not depend on a different synaptic organization of the rRTN.

Serotonin 5-HT1A receptors might control the output of cortical glutamatergic neurons in rat cingulate cortex

The present study was designed to investigate the distribution of serotonin 5-HT1A receptor protein (5-HT1A-immunoreactivity) and its localization within cortical pyramidal neurons of the rat cingulate cortex. This experimental direction was inspired by recent data showing the role of 5-HT1A receptors in the pathology of schizophrenia, and in the mechanism of action of novel antipsychotic drugs as well as by the importance of the cingulate cortex in regulation of cognitive functions. It was found that 5-HT1A-immunoreactivity was densely distributed in neuronal eyelash-like elements, and their size, shape and spatial orientation may suggest concentration of 5-HT1A-immunopositive material in the proximal fragments of axons of cortical neurons. Moreover, it was observed that these 5-HT1A-immunopositive fragments were present predominately on proximal fragments of axons of pyramidal neurons, which was evidenced by double labeling experiments using glutamate and non-phosphorylated neurofilament H as markers of the cortical pyramidal cells. The 5-HT1A receptor immunoreactivity was localized distally to the inhibitory GABAergic terminals of chandelier and basket cells surrounding the pyramidal cell bodies and occasionally surrounding short initial segment of axonal hillock of pyramidal neurons. These anatomical data indicate that 5-HT1A receptors might control the excitability and propagation of information transmitted by the pyramidal cells. Moreover, our results indicate that drugs operating via 5-HT1A receptors in the cingulate cortex might control from this level the release of glutamate in the subcortical structures. Finally, the 5-HT1A receptors present in the cingulate cortex, as demonstrated in the present study, may constitute an important target for drugs used to repair dysfunction of glutamate neurotransmission, which is observed for example in schizophrenia.

Action potentials in basal and oblique dendrites of rat neocortical pyramidal neurons.

Basal and oblique dendrites comprise ~2/3 of the total excitable membrane in the mammalian cerebral cortex, yet they have never been probed with glass electrodes, and therefore their electrical properties and overall impact on synaptic processing are unknown. In the present study, fast multi-site voltage-sensitive dye imaging combined with somatic recording was used to provide a detailed description of the membrane potential transients in basal and oblique dendrites of pyramidal neurons during single and trains of action potentials (APs). The optical method allowed simultaneous measurements from several dendrites in the visual field up to 200 microm from the soma, thus providing a unique report on how an AP invades the entire dendritic tree. In contrast to apical dendrites, basal and oblique branches: (1) impose very little amplitude and time course modulation on backpropagating APs; (2) are strongly invaded by the somatic spike even when somatic firing rates reach 40 Hz (activity-independent backpropagation); and (3) do not exhibit signs of a 'calcium shoulder' on the falling phase of the AP. A compartmental model incorporating AP peak latencies and half-widths obtained from the apical, oblique and basal dendrites indicates that the specific intracellular resistance (Ri) is less than 100 omicron cm. The combined experimental and modelling results also provide evidence that all synaptic locations along basal and oblique dendrites, situated within 200 microm from the soma, experience strong and near-simultaneous (latency < 1 ms) voltage transients during somatic firing. The cell body, axon hillock and basal dendritic compartments achieve unique synchronization during each AP. Therefore, with respect to a retrograde signal (AP), basal and proximal oblique dendrites should be considered as an integral part of the axo-somatic compartment.

Hyperpolarization-activated current Ih disconnects somatic and dendritic spike initiation zones in layer V pyramidal neurons.

Layer V pyramidal cells of the somatosensory cortex operate with two spike initiation zones. Subthreshold depolarizations are strongly attenuated along the apical dendrite linking the somatic and distal dendritic spike initiation zones. Sodium action potentials, on the other hand, are actively back-propagating from the axon hillock into the apical tuft. There they can interact with local excitatory input leading to the generation of calcium action potentials. We investigated if and how back-propagating sodium action potentials alone, without concomitant excitatory dendritic input, can initiate calcium action potentials in the distal dendrite. In acute slices of the rat somatosensory cortex, layer V pyramidal cells were studied under current-clamp with simultaneous recordings from the soma and the apical dendrite. A train of four somatic action potentials had to reach high frequencies to induce calcium action potentials in the dendrite ("critical frequency," CF approximately 100 Hz). Depolarization in the dendrite reduced the CF, while hyperpolarization increased it. The CF depended on the presence of the hyperpolarization-activated current Ih: blockade with 20 microM 4-(N-ethyl-N-phenylamino)-1,2-dimethyl-6-(methylamino) pyridinium chloride (ZD7288) reduced the CF to 68% of control. If the neurons were stimulated with noisy current injections, leading to in-vivo-like irregular spiking, no calcium action potentials were induced in the dendrite. However, after Ih channel blockade, calcium action potentials were frequently seen. These data suggest that Ih prevents initiation of the dendritic calcium action potential by proximal input alone. Dendritic calcium action potentials may therefore represent a unique signature for coincident somatic and dendritic activation.

Identification and localisation of the sodium channel alpha subunits carrying the Ttx-sensitive sodium current (INa) in the enteric nervous system of the guinea PIG intestine

The sodium channel protein consists of an alpha subuint in association with auxiliary beta subunits. The 5 neuronal alpha subnnit isoforms that carry the TTX-sensittve current are differentially expressed according to the neuronal types and have different functional properties, which result in distinct conductances in specific cells. Our work was designed in order to identify which alpha subunits carry the TTX-sensitive Na current in the enteric nervous system. Experiments were performed on fixed whole mount preparations of the myenteric and submucous plexuses of guinea pig (GP) small and large intestine using polydonal antibodies (ABs) raised against rat and/or human epitopes of the following alpha subunits : Navl.1, Nay1.2, Navl.3, Navl.6 and Nay1.7. Positive controls on non enteric nervous system were obtained for all ABs except that against Nay1.7. AB selectivity was verified using Western blots performed on rat and GP brain membrane preparations. Only NaV1.2 and Navl.3 alpha subunits were expressed in enteric neurons. NaV1.2 was expressed in the majority of the enteric neurons and immunoreactivity was located in the soma. Interestingly, it was highly expressed in a single axon hillock of the enteric sensory neurons. Some of the myenteric NaV1.2 immunoreactive non-sensory neurons are likely to be motor neurons that innervate the longitudinal muscle. Navl.3-fike immunoreactivity was observed only in descending filamentous neurons It was located in the soma, the long ramifying dendrites and the axon. Numerous smooth nerve fibers were also labeled. These fibres are extrinsic in origin since they enter the gut wall in large bundles running from the mesentery. Using Western blots performed on adult and new born rat and GP brain membrane preparations, the Navl.2 AB, raised against epitopes located in the I-I1 intracellular loop (M1 IL), stained a band at 230 kDa. Using rat brain membrane preparations, the Navl.3 AB, also raised against an epitope located in 1-II 1L, stained a band at 130 kDa. A band of similar low molecular weight was also stained using a Pan AB raised against the loop llI-IV. The expression of both the 230 and 130 kDa protein was age-dependent as previously described for Navl.2 and Navl 3 (Felts eta;. 1997). Our data strongly suggests that in enteric sensory neurons and in some motor neurons of the GP the TTX-sensitive INa is exclusively car'ned by the Navl.2 alpha subunit. This work is supported by GSK and CNRS.

Redistribution of Syntaxin mRNA in Neuronal Cell Bodies Regulates Protein Expression and Transport during Synapse Formation and Long-Term Synaptic Plasticity

Syntaxin has an important role in regulating vesicle docking and fusion essential for neurotransmitter release. Here, we demonstrate that the distribution of syntaxin mRNA in cell bodies of sensory neurons (SNs) of Aplysia maintained in cell culture is affected by synapse formation, synapse stabilization, and long-term facilitation (LTF) produced by 5-HT. The distribution of the mRNA in turn regulates expression and axonal transport of the protein. Syntaxin mRNA and protein accumulated at the axon hillock of SNs during the initial phase of synapse formation. Significant numbers of granules containing syntaxin were detected in the SN axon. When synaptic strength was stable, both mRNA and protein were targeted away from the axon hillock, and the number of syntaxin granules in the SN axon was reduced. Dramatic increases in mRNA and protein accumulation at the axon hillock and number of syntaxin granules in the SN axon were produced when cultures with stable connections were treated with 5-HT that evoked LTF. Anisomycin (protein synthesis inhibitor) or KT5720 (protein kinase A inhibitor) blocked LTF, accumulation of syntaxin mRNA and protein at the axon hillock, and the increase in syntaxin granules in SN axons. The results indicate that without significant effects on overall mRNA expression, both target interaction and 5-HT via activation of protein kinase A pathway regulate expression of syntaxin and its packaging for transport into axons by influencing the distribution of its mRNA in the SN cell body.

A Dynamic Dendritic Refractory Period Regulates Burst Discharge in the Electrosensory Lobe of Weakly Electric Fish

Na+-dependent spikes initiate in the soma or axon hillock region and actively backpropagate into the dendritic arbor of many central neurons. Inward currents underlying spike discharge are offset by outward K+ currents that repolarize a spike and establish a refractory period to temporarily prevent spike discharge. We show in a sensory neuron that somatic and dendritic K+ channels differentially control burst discharge by regulating the extent to which backpropagating dendritic spikes can re-excite the soma. During repetitive discharge a progressive broadening of dendritic spikes promotes a dynamic increase in dendritic spike refractory period. A leaky integrate-and-fire model shows that spike bursts are terminated when a decreasing somatic interspike interval and an increasing dendritic spike refractory period synergistically act to block backpropagation. The time required for the somatic interspike interval to intersect with dendritic refractory period determines burst frequency, a time that is regulated by somatic and dendritic spike repolarization. Thus, K+ channels involved in spike repolarization can efficiently control the pattern of spike output by establishing a soma-dendritic interaction that invokes dynamic shifts in dendritic spike properties.

A modelling study of locomotion-induced hyperpolarization of voltage threshold in cat lumbar motoneurones.

During fictive locomotion the excitability of adult cat lumbar motoneurones is increased by a reduction (a mean hyperpolarization of approximately 6.0 mV) of voltage threshold (Vth) for action potential (AP) initiation that is accompanied by only small changes in AP height and width. Further examination of the experimental data in the present study confirms that Vth lowering is present to a similar degree in both the hyperpolarized and depolarized portions of the locomotor step cycle. This indicates that Vth reduction is a modulation of motoneurone membrane currents throughout the locomotor state rather than being related to the phasic synaptic input within the locomotor cycle. Potential ionic mechanisms of this locomotor-state-dependent increase in excitability were examined using three five-compartment models of the motoneurone innervating slow, fast fatigue resistant and fast fatigable muscle fibres. Passive and active membrane conductances were set to produce input resistance, rheobase, afterhyperpolarization (AHP) and membrane time constant values similar to those measured in adult cat motoneurones in non-locomoting conditions. The parameters of 10 membrane conductances were then individually altered in an attempt to replicate the hyperpolarization of Vth that occurs in decerebrate cats during fictive locomotion. The goal was to find conductance changes that could produce a greater than 3 mV hyperpolarization of Vth with only small changes in AP height (< 3 mV) and width (< 1.2 ms). Vth reduction without large changes in AP shape could be produced either by increasing fast sodium current or by reducing delayed rectifier potassium current. The most effective Vth reductions were achieved by either increasing the conductance of fast sodium channels or by hyperpolarizing the voltage dependency of their activation. These changes were particularly effective when localized to the initial segment. Reducing the conductance of delayed rectifier channels or depolarizing their activation produced similar but smaller changes in Vth. Changes in current underlying the AHP, the persistent Na(+) current, three Ca(2+) currents, the "h" mixed cation current, the "A" potassium current and the leak current were either ineffective in reducing Vth or also produced gross changes in the AP. It is suggested that the increased excitability of motoneurones during locomotion could be readily accomplished by hyperpolarizing the voltage dependency of fast sodium channels in the axon hillock by a hitherto unknown neuromodulatory action.

Axonal Na + channels calling the shots

The neuronal action potential is one of the most important aspects of inter-neuronal communication and brain function. However, relatively little is known about the precise determinants of the action-potential threshold or its site of initiation. Using state-of-the-art electrophysiological recording techniques to study the biophysical mechanisms underlying action potentials, Colbert and Pan now shine new light onto this question [1xIon channel properties underlying axonal action potential initiation in pyramidal neurons. Colbert, C.M. and Pan, E. Nat. Neurosci. 2002; 5: 533–538Crossref | PubMedSee all References][1]. It was previously thought, owing to the high density of Na+ channels in the axon hillock, that action potentials initiate near the neuronal soma. However, experimental evidence for this potential mechanism comes predominantly from theoretical and labeling studies rather than from functional studies. Now Colbert and Pan report direct electrophysiological recordings from neocortical axons and propose that the lowest threshold for action-potential initiation is in the axon itself rather than in the axonal hillock.Using outside-out patch-clamp recordings from somatic, initial axon segment (<30 μm from the soma) and axonal (30–47 μm from the soma) neuronal areas, the authors observed qualitative differences in the currents evoked upon depolarization in these regions. In somatic patches, a combination of Na+ and delayed-rectifier K+ currents, as well as fast A-type K+ currents, could be evoked, whereas fast A-type K+ currents were absent in initial segment and axonal areas. In addition, it was observed that Na+ channels in the axon require significantly less depolarization for activation than do those in the soma or initial segment. This shift did not decrease gradually from soma to axon, but rather, changed sharply at the initial segment. Current densities were uniform throughout the initial segment before increasing two- to threefold in the axon.Using numerical models of neurons, and equivalent shifts in axonal Na+-channel activation, the authors found a decrease in axonal threshold for action-potential initiation with respect to the soma that corresponded well with the experimental data. However, modeling an increase in Na+-channel density in the initial segment only affected the axonal threshold at very high channel densities. In both situations, the site of initiation was biased away from the soma, although the shifted channel activation could better describe the experimental evidence for axonal action-potential initiation.This work provides a biophysical description of the axonal ion channels that might be involved in the initiation of action potentials. Axonal Na+ channels were found to require less depolarization for activation than did those in the soma or initial segment of the axon – this shift in activation kinetics could account for a lower threshold for action-potential initiation in the axon versus the soma. These data suggest that it is not the number of Na+ channels in the axon hillock but, rather, the properties of the Na+ channels in the axon that are responsible for the low axonal action-potential threshold. It is important not to overlook the role of other ion channels with respect to action-potential initiation and the diversity of neuronal physiologies; however, the data reported by Colbert and Pan in this study indicate that the likely determinants of neuronal action-potential initiation are located in the axon.