Comparative histomorphology of the Mauthner cells in some freshwater teleosts.

The histomorphological observations are made on the Mauthner cells in eight species of teleosts belonging to six different families. The cells are better developed in Channa punctatus, Heteropneustes fossilis, Labeo rohita, Danio, malabaricus and Puntius ticto. They are symmetrically situated in Nandus nandus and are found to be absent in Mastocembalus armatus. Their position, shape and size vary in different species. The axon cap is well developed in Channa punctatus, Heteropneustes fossilis and carps. The cell body sends lateral and ventral dendrites besides several small dendrites. The lateral dendrite emerges through the axon cap, turns dorsolateral and becomes myelinated to form Mauthner axon. The Mauthner axon extends in the spinal region upto the caudal peduncle and forms synapses with the spinal motoneurons of the front column. There are numerous synapses and end bulbs from the vestibular fibres and VIIIth nerve distributed on the perikaryan of the Mauthner cell body. It is suggested that the Mauthner cells are comparatively well developed in those species in which the tail fin is better utilized for swimming.     

Mauthner cell-initiated abrupt increase of the electric organ discharge in the weakly electric fishGymnotus carapo

Stimulation of the spinal cord of the electric fish Gymnotus carapo, evoked an abrupt increase in the discharge rate of the electric organ. At the maximum of this response, the rate increased an average of 26 ± 11.8%. The duration of the response was 4.9 ± 2.12 s; its latency was 10.4 ± 1.1 ms. Activation of the Mauthner axon played a decisive role in this phenomenon as indicated by the following: (1) recordings from the axon cap of the Mauthner cell demonstrated that the response was evoked if the Mauthner axon was antidromically activated and (2) a response that was similar to that produced by spinal cord stimulation, was elicited by intracellular stimulation of either Mauthner cell. Stimulation of the eighth nerve could also increase the discharge rate of the electric organ. The effect was greater if a Mauthner cell action potential was elicited. The findings described in the present report, indicate the existence of a functional connection between the Mauthner cell and the electromotor system in Gymnotus carapo. This connection may function to enhance the electrolocative sampling of the environment during Mauthner-cell mediated behaviors. This is a novel function for the Mauthner cell.Abbreviations EHP extrinsic hyperpolarizing potential - EOD electric organ discharge - M-AIR Mauthner initiated abrupt increase in rate - M-cell Mauthner cell - M-axon Mauthner axon - PM pacemaker nucleus - PM-cell pacemaker cell - PPn prepacemaker nucleus - SPPn sublemniscal prepacemaker nucleus     

Mauthner cell-evoked synaptic actions on pacemaker medullary neurons of a weakly electric fish

The present study was designed to examine the synaptic events in neurons of the pacemaker nucleus of Gymnotus carapo during the increase in rate of the electric organ discharge following activation of Mauthner cells. Pacemaker and relay cells were investigated using intracellular recordings which were performed under two different conditions: (1) with the pacemaker nucleus spontaneously discharging and (2) after its activity was abolished by anesthesia. Mauthner axon activation induced an increase in the rate of pacemaker cell discharges. This response was accompanied by an increase in the slope of the pacemaker potential (up to 110%) and a depolarization of these cells. The discharges of relay cells followed one to one those of pacemaker cells. In contrast to that observed in pacemaker cells, only brief depolarizing antidromic effects could be evoked in relay cells after Mauthner axon activation. In quiescent pacemaker cells, Mauthner cell activation induced a prolonged (up to 500 ms) depolarizing potential with an average amplitude of 1.92 ± 0.82 mV; its latency was 4.43 ± 1.14 ms. Our data indicate that, within the pacemaker nucleus, the population of pacemaker cells is the only target for Mauthner cell-evoked, short-latency excitatory synaptic actions. Accepted: 1 March 1997     

Development of the axon cap neuropil of the Mauthner cell in the goldfish

Summary Development of the axon cap neuropil of the Mauthner neuron in post-hatching larval goldfish brains was observed electron-microscopically. The axonal initial segment of newly hatched (day-4) larvae is completely covered with synaptic terminals containing clear spherical synaptic vesicles. Profiles of thin terminal axons, the spiral fibers, containing similar synaptic vesicles, rapidly increase in number around the initial segment and form glomerular neuropil similar to the central core of the adult axon cap by day 7. Three types of synapses are formed in the core neuropil. Bouton-type synapses contacting the initial segment are most abundant in day-4 to-14 larvae; they decrease thereafter and are rare on the distal half of the initial segment of day-40 larvae. Asymmetric axo-axonic synapses are commonly observed between spiral fibers in the core neuropil of day-7 to -19 larvae, but become fewer by day 40. Unique symmetrical axo-axonic synapses showing accumulation of synaptic vesicles on either side of apposed membrane thickenings first appear in day-14 core neuropil, gradually increase in number, and become the predominant type in day-40 core neuropil. Thick myelinated axons, which lose their myelin sheaths in the glial cap cell layer, start to penetrate into the axon cap on day 10. They gradually increase in number and form the peripheral part of the axon cap together with the cap dendrites, which finally grow into the axon cap from the axon hillock region of the Mauthner cell by day 40.     

PROTEIN SYNTHESIS IN THE ISOLATED MAUTHNER NERVE FIBRE OF GOLDFISH

Abstract— Mauthner nerve fibres isolated from the spinal cord of goldfish were incubated, in the presence of radioactive amino acids for varying periods of time. It was found that the Mauthner fibre synthesizes proteins in the absence of cell nuclei. Amino acid incorporation showed sensitivity to puromycin and to acetoxycycloheximide but resistance to chloramphenicol. Only slight inhibition was caused by actinomycin-D. The contribution of the denuded axon to the total protein synthesis was about 30 per cent per unit length Mauthner fibre. The remaining activity was due to the myelin sheath compartment. Fractionation experiments showed that the incorporation in the sheath was due to components other than the myelin lamellae. The subcellular distribution of newly synthesized proteins in the isolated and incubated Mauthner fibre was compared to that found in the incubated spinal cord. The results strongly suggested the existence in the Mauthner fibre of a primary microsomal, rather than a mitochondrial, protein synthesizing system.     

The reaction of endoplasmic reticulum of Mauthner cells of Xenopus laevis tadpoles in response to the partial denervation

The structure of the Mauthner cells in Xenopus laevis tadpole was investigated by light- and electron microscopy in norm and after early unilateral enucleation. It was found that enucleation at early stages caused a delay in morphological development of the contralateral neurons during embryogenesis. We observed a decrease in size of the soma and nucleus and in the number of dendrites, a marked structural underdevelopment of the majority of cell organoids, as well as proliferation and hypertrophy of transversal cisternae in the contralateral Mauthner cells. The ipsilateral neurostructure remained normal in embryogenesis. The data obtained suggest the availability of some unknown powerful afferent contralateral input to Mauthner cells from the optical analyzer.     

Individual axons regulate the myelinating potential of single oligodendrocytes in vivo

The majority of axons in the central nervous system (CNS) are eventually myelinated by oligodendrocytes, but whether the timing and extent of myelination in vivo reflect intrinsic properties of oligodendrocytes, or are regulated by axons, remains undetermined. Here, we use zebrafish to study CNS myelination at single-cell resolution in vivo. We show that the large caliber Mauthner axon is the first to be myelinated (shortly before axons of smaller caliber) and that the presence of supernumerary large caliber Mauthner axons can profoundly affect myelination by single oligodendrocytes. Oligodendrocytes that typically myelinate just one Mauthner axon in wild type can myelinate multiple supernumerary Mauthner axons. Furthermore, oligodendrocytes that exclusively myelinate numerous smaller caliber axons in wild type can readily myelinate small caliber axons in addition to the much larger caliber supernumerary Mauthner axons. These data indicate that single oligodendrocytes can myelinate diverse axons and that their myelinating potential is actively regulated by individual axons.     

SEDIMENTATION ANALYSIS OF RIBONUCLEIC ACID EXTRACTED FROM ISOLATED MAUTHNER NERVE FIBRE COMPONENTS

Abstract— Segments of spinal cords from goldfish or from carp were incubated in vitro in the presence of RNA precursors for varying periods of time. Mauthner nerve fibres were isolated from the fresh unfixed tissue, or, for the separate analyses of axon and myelin sheath, from the fixed spinal cord.
The myelinated Mauthner fibre isolated from the incubated spinal cord showed RNA synthesis. A considerable part of the material sedimented at 4 S , but part of the nucleic acid was recovered at higher sedimentation values, up to 30 S. Newly synthesized RNA was extracted from the isolated myelin sheath as well as from the axon. Isolated myelinated Mauthner fibres were also incubated in vitro with RNA precursors. In this case incorporation occurred exclusively in material sedimenting at 4 S or lower. The turnover rate for RNA from the fibre was of a higher order than that of the bulk of RNA from the spinal cord. The findings of RNA synthesis in these tissue components lacking nuclei could possibly be explained as owing to mitochondria.
Studies by electron microscopy demonstrated the extent of purity of the isolated components and it was found that contamination was so small as to make it unlikely that the RNA investigated originated in contaminating tissue.     

Zebrafish deadly seven functions in neurogenesis.

In a genetic screen, we isolated a mutation that perturbed motor axon outgrowth, neurogenesis, and somitogenesis. Complementation tests revealed that this mutation is an allele of deadly seven (des). By creating genetic mosaics, we demonstrate that the motor axon defect is non-cell autonomous. In addition, we show that the pattern of migration for some neural crest cell populations is aberrant and crest-derived dorsal root ganglion neurons are misplaced. Furthermore, our analysis reveals that des mutant embryos exhibit a neurogenic phenotype. We find an increase in the number of primary motoneurons and in the number of three hindbrain reticulospinal neurons: Mauthner cells, RoL2 cells, and MiD3cm cells. We also find that the number of Rohon-Beard sensory neurons is decreased whereas neural crest-derived dorsal root ganglion neurons are increased in number supporting a previous hypothesis that Rohon-Beard neurons and neural crest form an equivalence group during development. Mutations in genes involved in Notch-Delta signaling result in defects in somitogenesis and neurogenesis. We found that overexpressing an activated form of Notch decreased the number of Mauthner cells in des mutants indicating that des functions via the Notch-Delta signaling pathway to control the production of specific cell types within the central and peripheral nervous systems.     

Stereotyped and variable growth of redirected Mauthner axons

To assay the axon tract organizing capabilities of different regions of the vertebrate CNS, Mauthner axons were redirected by grafting supernumerary hindbrains in Xenopus embryos. The 63 redirected Mauthner axons thus produced included donor axons projecting into the host CNS and host axons that grew through the graft or that were redirected in the host CNS. Two major phenomena were observed. Caudal to the optic chiasm, the Mauthner axons followed a single ipsilateral stereotyped route—the basal substrate pathway—extending in the ventral and ventrolateral marginal zone from the diencephalon to the caudal spinal cord. In contrast, rostral to the optic chiasm, these same Mauthner axons followed variable ipsilateral and contralateral routes. Even pairs of Mauthner axons entering the optic chiasm side-by-side eventually followed different routes in normal forebrains. The contrasting behaviors of the Mauthner axons growing in the rostral diencephalon and telencephalon and of the same Mauthner axons growing elsewhere suggest that there are differences in the effective guidance cues between these two regions of the developing brain. This is consistent with other types of neuroanatomical and neuroembryological evidence indicating a fundamental division between the rostral and the caudal diencephalon.     

Invasion of microglia/macrophages and granulocytes into the Mauthner axon myelin sheath following spinal cord injury of the adult goldfish,Carassius auratus

Rapid activation of resident glia occurs after spinal cord injury. Somewhat later, innate and adaptive immune responses occur with the invasion of peripheral immune cells into the wound site. The activation of resident and peripheral immune cells has been postulated to play harmful as well as beneficial roles in the regenerative process. Mauthner cells, large identifiable neurons located in the hindbrain of most fish and amphibians, provided the opportunity to study the morphological relationship between reactive cells and Mauthner axons (M-axons) severed by spinal cord crush or by selective axotomy. After crossing in the hindbrain, the M-axons of adult goldfish, Carassius auratus, extend the length of the spinal cord. Following injury, the M-axon undergoes retrograde degeneration within its myelin sheath creating an axon-free zone (proximal dieback zone). Reactive cells invade the wound site, enter the axon-free dieback zone and are observed in the vicinity of the retracted M-axon tip as early as 3 hr postinjury. Transmission electron microscopy allowed the detection of microglia/macrophages and granulocytes, some of which appear to be neutrophil-like, at each of these locations. We believe that this is the first report of the invasion of such cells within the myelin sheath of an identifiable axon in the vertebrate central nervous system (CNS). We speculate that microglia/macrophages and granulocytes that are attracted within a few hours to the damaged M-axon are part of an inflammatory response that allows phagocytosis of debris and plays a role in the regenerative process. Our results provide the baseline from which to utilize immunohistochemical and genetic approaches to elucidate the role of non-neuronal cells in the regenerative process of a single axon in the vertebrate CNS.     

Decrease in occurrence of fast startle responses after selective Mauthner cell ablation in goldfish (Carassius auratus )

A single action potential in one of a pair of reticulospinal neurons, the Mauthner cells, precedes a short-latency electromyographic response of the trunk and tail musculature on the opposite side of the body and a fast startle response in goldfish. It has been postulated that not only the Mauthner cell, but also an array of neurons can trigger or participate in fast startle responses (Eaton et al. 1991). We have selectively ablated the Mauthner cells in goldfish to study how neurons of the brainstem fast startle response network interact. The probability of eliciting a fast startle response was significantly less in fish with double Mauthner cell ablations, as compared to the responsiveness of control fish. The finding that there is a significant decrease in the occurrence of fast startle responses in animals with no Mauthner cells, implies that the Mauthner cell may play a role in triggering the involvement of the other network elements in fast startle responses. We hypothesize that Mauthner cell activation may be important in bringing those reticulospinal neurons that are “primed” by the behavioral context to threshold and provides the basis for studies focused on the interactive nature of the brainstem startle response network. Accepted: 4 November 1998     

Bergmann glia and the recognition molecule CHL1 organize GABAergic axons and direct innervation of Purkinje cell dendrites

The geometric and subcellular organization of axon arbors distributes and regulates electrical signaling in neurons and networks, but the underlying mechanisms have remained elusive. In rodent cerebellar cortex, stellate interneurons elaborate characteristic axon arbors that selectively innervate Purkinje cell dendrites and likely regulate dendritic integration. We used GFP BAC transgenic reporter mice to examine the cellular processes and molecular mechanisms underlying the development of stellate cell axons and their innervation pattern. We show that stellate axons are organized and guided towards Purkinje cell dendrites by an intermediate scaffold of Bergmann glial (BG) fibers. The L1 family immunoglobulin protein Close Homologue of L1 (CHL1) is localized to apical BG fibers and stellate cells during the development of stellate axon arbors. In the absence of CHL1, stellate axons deviate from BG fibers and show aberrant branching and orientation. Furthermore, synapse formation between aberrant stellate axons and Purkinje dendrites is reduced and cannot be maintained, leading to progressive atrophy of axon terminals. These results establish BG fibers as a guiding scaffold and CHL1 a molecular signal in the organization of stellate axon arbors and in directing their dendritic innervation.     

Phosphorylation of Neurofilament Proteins in Isolated Goldfish Mauthner Axoplasm

Abstract: The six neurofilament proteins (NFPs) in the goldfish Mauthner axon (M-axon) have molecular sizes of 235, 145, 123, 105, 80, and 60 kDa. To determine if NFPs in the M-axon are phosphorylated, isolated Mauthner axoplasm (M-axoplasm) and a neurofilament-enriched extract (NFE) prepared from M-axoplasm were incubated with ³²P, which resulted in the radiolabeling of NFPs as determined by their detection on autoradiograms. Kinase inhibitors directed against cyclic AMP-dependent kinases (PKAs) or cofactor-independent kinases significantly reduced the in vitro phosphorylation of NFPs in NFE, whereas inhibitors directed against protein kinase C did not significantly reduce the in vitro phosphorylation of NFPs in NFE. Experiments using two kinase inhibitors directed against different kinases significantly reduced the in vitro phosphorylation of NFPs in NFE to a greater extent than the reduction produced using any single kinase inhibitor. These data suggest that NFPs in the M-axon are phosphorylated and that the in vitro (and perhaps the in vivo) phosphorylation of NFPs is mediated by PKA and/or cofactor-independent kinases that copurify with NFPs.     

AN ELECTRON MICROSCOPIC CHARACTERIZATION OF CLASSES OF SYNAPTIC VESICLES BY MEANS OF CONTROLLED ALDEHYDE FIXATION

Examination of variables of aldehyde fixation that may affect the shape of agranular synaptic vesicles has revealed that even brief storage of aldehyde-perfused nervous tissue pieces in cacodylate buffer, prior to hardening in osmium tetroxide, has an unusually severe flattening effect on agranular vesicles of a particular type. These are the vesicles of peripheral cholinergic axon endings, and of certain central synaptic bulbs. Types of synaptic bulbs can now be further defined on the basis of shape of agranular synaptic vesicles under controlled conditions of aldehyde fixation. Previously described "S" bulbs in the spinal cord contain uniformly spheroid vesicles, which are wholly resistant to flattening. Previously described "F" bulbs contain somewhat smaller agranular vesicles that are flattened after aldehyde fixation, even when this is followed by prompt hardening in osmium tetroxide solution. A third type, previously characterized as having irregularly round agranular vesicles after the above treatment, contains only severely flattened vesicles when the osmium tetroxide hardening is preceded by even a brief wash with sodium cacodylate buffer containing sucrose. Moreover, the "third" type is characteristic of all cholinergic peripheral axon endings examined, as well as the large axosomatic ("L") synaptic bulbs of the spinal cord.     

刺激鲫鱼小脑腹外侧区诱发的Mauthner细胞兴奋性突触后电位

实验采用微电极胞内记录技术探查鲫鱼Mauthner细胞(M-细胞)对小脑刺激的电反应特征。电刺激鲫鱼小脑腹外侧部,可在双侧M-细胞胞体、腹侧树突和外侧树突近端记录到一种复合性兴奋性突触后电位(小脑诱发性EPSP)。小脑诱发性EPSP潜伏期较短(0.63±0.09 ms),持续时间较长(5.49±1.13 ms),幅度分级和刺激频率依从等特征。以较高强度刺激小脑常引起M-细胞顺向激活。多点胞内连续穿刺实验显示小脑诱发性EPSP起源于腹侧树突远端。实验结果提示,小脑-M-细胞通路可能包含一组长短不等的神经元链,它们根据链的短或长,由近及远依次投射在腹侧树突远端。     

The Neuroprotective Effect of the Thr–Ser–Lys–Tyr Peptide in a Goldfish Mauthner Cell Model in vivo

The effects of the Thr–Ser–Lys–Tyr peptide, which was shown to display neuroprotective activity in cell cultures in vitro, were studied in the model of paired Mauthner cells of goldfish. It was found that intracerebral injections provided the peptide to be applied into the zone of the right Mauthner cell under the fourth ventricle of the hindbrain lead to a dose-dependent decrease in the number of spontaneous turns of the goldfish to the left. It was shown that this effect is not eliminated under long-lasting optokinetic stimulation when the fish instinctively follow stimuli with a low spatial frequency that are moving in the nasal-to-temporal direction. We used the method of three-dimensional reconstruction by serial histological sections to study the dendrite morphology of the Mauthner cells in control and experimental goldfish. It was found that optokinetic stimulation of control goldfish evokes the dystrophy of the ventral dendrite of the right Mauthner cell, which is the target of this type of stimulation. Conversely, the peptide stabilize the size of the ventral dendrite of the right Mauthner cell under stimulation. These data could be interpreted as evidence of the neuroprotective effect of the Thr–Ser–Lys–Tyr peptide in vivo.     

THE ULTRASTRUCTURE OF MAUTHNER CELL SYNAPSES AND NODES IN GOLDFISH BRAINS

An electron microscope study of goldfish Mauthner cells is reported.¹ The cell is covered by a synaptic bed ~ 5 µ thick containing unusual amounts of extracellular matrix material in which synapses and clear glia processes are implanted. The preterminal synaptic neurites are closely invested by an interwoven layer of filament-containing satellite cell processes. The axoplasm of the club endings contains oriented mitochondria, neurofilaments, neurotubules, and relatively few synaptic vesicles. That of the boutons terminaux contains many unoriented mitochondria and is packed with synaptic vesicles and some glycogen but no neurofilaments or neurotubules. The bare axons of club endings are surrounded by a moderately abundant layer of matrix material. The synaptic membrane complex (SMC) in cross-section shows segments of closure of the synaptic cleft ~ 0.2 to 0.5 µ long. These alternate with desmosome-like regions of about the same length in which the gap widens to ~ 150 A and contains a condensed central stratum of dense material. Here, there are also accumulations of dense material in pre- and postsynaptic neuroplasm. The boutons show no such differentiation and the extracellular matrix is largely excluded around them. The axon cap is a dense neuropil of interwoven neural and glial elements free of myelin. It is covered by a closely packed layer of glia cells. The findings are interpreted as suggestive of electrical transmission in the club endings.     

Potentiation of electrical and chemical synaptic transmission mediated by endocannabinoids

Endocannabinoids are well established as inhibitors of chemical synaptic transmission via presynaptic activation of the cannabinoid type 1 receptor (CB1R). Contrasting this notion, we show that dendritic release of endocannabinoids mediates potentiation of synaptic transmission at mixed (electrical and chemical) synaptic contacts on the goldfish Mauthner cell. Remarkably, the observed enhancement was not restricted to the glutamatergic component of the synaptic response but also included a parallel increase in electrical transmission. This effect involved the activation of CB1 receptors and was indirectly mediated via the release of dopamine from nearby varicosities, which in turn led to potentiation of the synaptic response via a cAMP-dependent protein kinase-mediated postsynaptic mechanism. Thus, endocannabinoid release can potentiate synaptic transmission, and its functional roles include the regulation of gap junction-mediated electrical synapses. Similar interactions between endocannabinoid and dopaminergic systems may be widespread and potentially relevant for the motor and rewarding effects of cannabis derivatives.