METABOLIC AND CHEMICAL CHANGES IN REGENERATING NEURONS—II <i>IN VITRO</i> RATE OF INCORPORATION OF AMINO ACIDS INTO PROTEINS OF THE NERVE CELL PERIKARYON OF THE C.8 SPINAL GANGLION OF RABBIT

Miani, N.; Rizzoli, Antonio A.; Bucciante, G

doi:10.1111/j.1471-4159.1961.tb13500.x

Cited by 34 publications

(4 citation statements)

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“…Murray and Grafstein (1) presented radioautographic evidence that the morphological changes were accompanied by increased amino-acid incorporation into retinal ganglion cell protein during regeneration of the optic nerve. Studies on mammalian cranial and spinal motor neurons (2)(3)(4) and on mammalian spinal ganglion cells (5,6) have also been interpreted to indicate increased protein synthesis in these neuronal perikarya following axotomy. The increased incorporation of amino acid into protein of the nerve cell body is accompanied by an increased rate of axonal transport in the regenerating axon (7)(8)(9).…”

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Enhanced labeling of a retinal protein during regeneration of optic nerve in goldfish.

Heacock¹,

Agranoff²

1976

Proc. Natl. Acad. Sci. U.S.A.

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Goldfish retinas were examined for changes in the labeling pattern of protein during regeneration of retinal ganglion cell axons following unilateral optic nerve crush. At various times after optic nerve crush the normal retinas were incubated in vitro with [3H] Following intra-orbital crush of the optic nerve, in goldfish, the retinal ganglion cell must grow out a new axon several millimeters long. This process not only implies a substantial increase in the rate of protein synthesis but may also involve a selective increase in the synthesis of the proteins necessary for the renewal of axoplasm and replacement of structural elements of the axon. Formation of cell-specific markers involved in retinotectal recognition may be increased~during this period (10). There might, in addition, be an initial decrease in the synthesis of components necessary to maintain the functional integrity of the nerve ending until the growing nerve reaches the optic tectum (11).This report presents comparisons of the in vitro protein labeling pattern of normal goldfish retina with that of retinas removed at various times after optic nerve crush (postcrush retina). A double-labeling technique coupled with sodium dodecyl sulfate-urea polyacrylamide gel electrophoresis (12) revealed an increased amino-acid incorporation into a protein fraction with properties of the microtubule subunit, tubulin. MATERIALS AND METHODSGoldfish (Carassius auratus), 6-7 cm in body length, were anesthetized with tricaine methane sulfonate prior to an intraorbital crush of the right optic nerve. The left optic nerve remained intact so that the left retina served as a control. Following surgery, the goldfish were stored in groups of 50 to 80 in 30 gallon tanks at 20-22°, and were fed daily.Fish were dark-adapted for 30 min prior to removal of retinas in order to facilitate separation of the retina from the pigment epithelium. After hemisection of the eye, the retina was floated free from the pigment layer with 0.86% saline and detached by cutting at the optic disk. The retinas were then rinsed and stored in ice-cold saline or incubation medium for up to 1 hr prior to incubation.In were evaporated to dryness under nitrogen immediately before use. After evaporation, each was resuspended in a small volume of incubation medium. The purity of each isotopic preparation was checked by paper chromatography. The incubation medium was that of Dunlop et al. (13): N-2-bydroxyethylpiperazine-N'-2-ethanesulfonic acid (Hepes), 25 mM; MgSO4, 1.3 mM; CaCl2, 2.6 mM; K2HPO4, 1.2 mM; KCI, 5.9 mM; NaCl, 106.5 mM; glucose, 12 mM; and NaOH to pH 7.4.Retinas (in groups of 5 to 20) were pre-incubated for 2 min at 25°in 1.9 ml of incubation medium. Labeled precursor (0.1 ml) was then added (1.0-4.0 ,uCi per retina) and incubation was continued in air in a Dubnoff metabolic shaker at 250. All incubations using double-labeled methionine were carried out for 1 hr. The reaction was stopped by dilution with 2 volumes of ice-cold incubation medium containing 2 mM L-methionine. After c...

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confidence: 99%

Enhanced labeling of a retinal protein during regeneration of optic nerve in goldfish.

Heacock¹,

Agranoff²

1976

Proc. Natl. Acad. Sci. U.S.A.

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“…Because the synthetic machinery of the regenerating neurons could operate presumably at near maximal capacity, a portion of the engulfed material might be recycled, or the processed elementary constituents reutilized by the neurons to meet the increased metabolic demands for reconstitution of cell membranes requested after axonal injury. In addition, it has been shown that regeneration of the peripheral processes of the unipolar cells of the C8 spinal ganglion in the rabbit occurs without changes in synthesis of phosphatidylcholine, ethanolamine, and serine plasmalogens, although a temporary increase of phosphatidylethanolamine and phosphatidylinositol was detected (Miani, 1962). The same interpretation could not be correct for the cervicothoracic system in which many of the axoplasm and axonal processes were missing after nerve crush in the vicinity of the cell bodies, and for which data on the changes of axonal transport of phospholipids during regeneration are lacking.…”

Section: Discussionmentioning

confidence: 99%

Changes of Phospholipid‐Metabolizing and Lysosomal Enzymes in Hypoglossal Nucleus and Ventral Horn Motoneurons During Regeneration of Craniospinal Nerves

Alberghina

Stella

1988

Journal of Neurochemistry

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In order to study the biochemical changes associated with the cell body response to axonal crush injury, two systems, hypoglossal nucleus and spinal cord ventral horn, were used. The time intervals chosen were 7, 14, and 28 days after unilateral crushing of the right hypoglossal nerve and cervicothoracic nerves of the rabbit. Non-crushed, contralateral nerves were used as controls. Three groups of enzyme activities were tested: (a) phospholipase A2, acyl CoA:2-acyl-sn-glycero-3-phosphocholine acyltransferase, and choline phosphotransferase, as indicators of phospholipid degradation and biosynthesis; (b) seven hydrolases, namely, beta-D-glucuronidase, beta-N-acetyl-D-hexosaminidase, arylsulfatase A, galactosylceramidase, GM1-ganglioside beta-galactosidase, and acid RNase, as indicators of lysosomal activity; and (c) free and inhibitor-bound alkaline RNase, as an index of RNA metabolism. Changes could be grouped into three distinct patterns. Compared to contralateral control, choline phosphotransferase showed a slight increase, whereas phospholipase A2 and most lysosomal hydrolases showed a significant increase of activity, especially evident in the ventral spinal cord neurons 14-28 days after crushing. These changes correlate with known increases of membrane and organelle numbers, including lysosomes, in motor and sensory neurons during peripheral regeneration. In contrast, free and acid alkaline RNase activity significantly decreased in the injured sides compared to the controls. This change can probably be correlated with a stabilization of RNAs needed for increased protein synthesis. No changes in total alkaline RNase and acyltransferase activities in either regeneration model were observed.

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“…An increased protein synthesis as indicated by increased incorporation of labelled amino acids into proteins in motoneurones and dorsal root 258 S. OCHS TRANSPORT IN CHROMATOLYSED NEURONES ganglia during chromatolysis (Miani, Rizzoli & Bucciante, 1961;Francoeur & Olszewski, 1969) or an apparently decreased level of synthesis by a lowered level of incorporation (Engh et al 1971) have both been reported. Possibly variations in both synthesis and proteolysis (Watson, 1965) may account for such different results.…”

Section: Amounts Of Incorporated Materials In Crests Of Transected Anmentioning

confidence: 99%

Fast axoplasmic transport in the fibres of chromatolysed neurones.

Ochs¹

1976

The Journal of Physiology

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3. The amount of transport shown by the level of activity in the crests on the chromatolytic and control sides relative to the 'pool' of radioactive materials remaining in the cell bodies of the ganglion were also similar. The significance of these findings was discussed with respect to changes in the cell bodies known to take place during chromatolysis and the stability of the axoplasmic transport mechanism in nerve fibres.

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METABOLIC AND CHEMICAL CHANGES IN REGENERATING NEURONS—II IN VITRO RATE OF INCORPORATION OF AMINO ACIDS INTO PROTEINS OF THE NERVE CELL PERIKARYON OF THE C.8 SPINAL GANGLION OF RABBIT

Cited by 34 publications

References 25 publications

Enhanced labeling of a retinal protein during regeneration of optic nerve in goldfish.

Enhanced labeling of a retinal protein during regeneration of optic nerve in goldfish.

Changes of Phospholipid‐Metabolizing and Lysosomal Enzymes in Hypoglossal Nucleus and Ventral Horn Motoneurons During Regeneration of Craniospinal Nerves

Fast axoplasmic transport in the fibres of chromatolysed neurones.

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