Light and electron microscope studies were conducted on the nature of the degenerative changes in amputated nerve fibers of cultured rat sensory ganglia and on the effects of media with differing calcium concentrations upon these changes. With glucose-enriched Eagle's media (MEM) containing 1.6 mM calcium, the amputated myelinated and unmyelinated axons undergo a progressive granular disintegration of their axoplasm with collapse and fragmentation of myelin sheaths between 6 and 24 h after transection. With MEM containing only 25–50 µM calcium, the granular axoplasmic degeneration does not occur in transected fibers and they retain their longitudinal continuity and segmental myelin ensheathment for at least 48 h. Addition of 6 mM EGTA to MEM (reducing the estimated Ca++ below 0.3 µM) results in the structural preservation of both microtubules and neurofilaments within transected axons. A transient focal swelling of amputated axons occurs, however, in cultures with normal and reduced calcium. These observations suggest that an alteration in the permeability of the axolemma is a crucial initiating event leading to axonal degenerative changes distal to nerve transection. The loss of microtubules and neurofilaments and the associated granular alterations of the axoplasm in transected fibers appears to result from the influx of calcium into the axoplasm.

https://rupress.org/jcb/article-pdf/59/2/456/1386704/456.pdf

Effects of calcium ion concentration on the degeneration of amputated axons in tissue culture.

A non-invasive blood analyte concentration monitor. The monitor includes a stimulator applying a stimulus to an endogenous tissue responsive to the stimulus and a detector for detecting a response of an endogenous tissue to the stimulus. Also provided is a correlator which correlates the detected response of the endogenous tissue to an analyte concentration. In one embodiment, the monitored blood analyte is glucose.

Apparatus and method for non-invasive blood analyte measurement

The authors have agreed that this Chapter should not be an exhaustive compilation of all papers. Instead they intend to submit the essential modern knowledge necessary to neurobiologists working with peripheral nerve of frogs which, in their opinion, is an essential test object to develop the theoretical and experimental basis of the function of excitable membranes of vertebrates. They want, in particular, to present a chapter which in the future can be used to familiarize newcomers with the methods and theories used in this field. Since there is almost no work on unmyelinated amphibian fibers, the discussion is almost exclusively on myelinated nerve.

Electrophysiology of the Peripheral Myelinated Nerve

The cardinal electrophysiological abnormalities in experimental diabetic (EDN) and experimental galactose (EGN) neuropathy, models in which endoneurial hypoxia has been demonstrated, are a slowing in nerve conduction velocity (NCV) and a resistance to ischemic conduction block (RICB). These electrophysiological abnormalities are also present in human diabetic neuropathy, where microangiopathy has been demonstrated to be more severe than in EDN so that endoneurial hypoxia is also likely to be present. We examined the effects of endoneurial hypoxia per se on normal nerves. In rats subjected to chronic hypoxia (10% O2) for up to 10 wk, the two electrophysiological abnormalities had developed by 4 wk and were very similar in degree to those seen in EDN and EGN. These abnormalities occurred in the absence of hyperglycemia, nerve sorbitol accumulation, or myoinositol reduction. Resting O2 consumption was reduced, the percent increase in nerve lactate under anoxic stress was increased, and nerve free sugars were normal. Hexokinase and phosphofructokinase activities were not altered substantially when studied under conditions of O2 excess. These findings indicate that hypoxia per se will cause conduction slowing and suggest that the hypoxic nerve develops RICB because of a reduced energy requirement and an increased efficiency of anaerobic glycolysis, but without major changes in the activity of its controlling glycolytic enzymes.

Experimental chronic hypoxic neuropathy: relevance to diabetic neuropathy.

Recent advances in the understanding of the pathogenesis of diabetic neuropathy have been made in six areas. (1) There is support for the notion that a reduction in nerve free myoinositol may be responsible in part for the nerve conduction slowing in diabetic neuropathy. (2) There is further evidence of microvascular abnormalities, including morphometric evidence of multifocal fiber loss and of capillary changes in biopsied sural nerve. (3) There is evidence of endoneurial hypoxia, including the findings of reduced nerve blood flow and endoneurial oxygen tensions in chronic experimental diabetic neuropathy (EDN). (4) The major mechanisms of resistance to ischemic conduction failure (RICF) is the marked increase in nerve energy substrates. (5) Recent studies provide certain insights into clinical characteristics of human diabetic neuropathy (HDN), including the assymetric pattern of HDN, the paradox between liability to pressure palsies and RICF, and insulin-related acute painful neuropathy. (6) The suggested pathogenetic scheme incorporates the notion that once hypoxia is established, it may start a vicious cycle of further capillary damage and escalating hypoxia.

Recent advances in the pathogenesis of diabetic neuropathy. Clues from human diabetic neuropathy

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Frog (Rana pipiens) sciatic nerve was incubated, with and without stimulation, in an oil bath. The correlation between changes in the magnitude of the compound action potential (α and β) and changes in metabolites, particularly energy reserves, during anoxia and recovery from anoxia was studied.



The time to extinction of the action potential in anoxia was frequency-dependent. The action potential could not be restored, nor its extinction delayed, by washing the nerve in O2-free Ringer's solution. Therefore, in this system extracellular K+ accumulation was not a significant factor in blocking impulse conduction.



At the time of complete nerve block resulting from anoxia (90 min at rest), ATP, P-creatine and glucose were 30, 10 and 10 per cent, respectively, of initial levels. Glycogen did not fall below 42 per cent of control levels even after 5 h of anoxia. Changes in the levels of energy reserves during anoxia were used to calculate the metabolic rate of nerves at rest and during stimulation. In one series of experiments, the resting metabolic rate was 0·12 mequiv. of ‘high-energy phosphate’ (∼P)/kg/min. Stimulation increased the metabolic rate to 0·22 mequiv. of ∼P/kg/min at 30 Hz and to 0·29 mequiv. of ∼P/kg/min at 100 Hz. The change in metabolic rate when the nerve passed from the resting to the stimulated state was quite abrupt, an observation suggesting that the slow transition observed with methods monitoring O2, consumption was largely instrumental.



In nerve stimulated to exhaustion in the absence of O2, neither ATP nor P-creatine had fully recovered within 60 min after O2, was readmitted, although the action potential reached supranormal levels 15 min after return to O2. The ratio of lactate: pyruvate, which increased as expected during anoxia, paradoxically increased even further after O2, was readmitted. The rate of energy utilization during recovery was 0·30 mequiv. of ∼P/kg/min.



Nerves stimulated at 100–200 Hz in O2, exhibited no changes in levels of P-creatine, ATP or lactate, an observation implying that the nerve could not be made to use ∼P faster than oxidation of glucose could provide it. This meant that the maximal metabolic rate was not limited by the rate of supply of chemical energy. Instead, the limitation may have arisen as a result of a limited rate at which ionic imbalance can result from stimulation or a limited pump capacity of the axonal membrane. Nerves stimulated at 200 Hz in O2 for 20 min and then transferred to an O2-free environment without further stimulation exhibited an increase in the rate of energy utilization (nearly two-fold) over the resting rate, a finding that suggested a metabolic (ionic?) debt as a result of activity which could not be met even though the energy supply was adequate. Therefore, restriction of energy expenditure by a limiting pumping rate seemed to be the most likely explanation.



The resting metabolic rate of frog sciatic nerve was only one-quarter to one-third of the rate for rat sciatic nerve, when compared at the same temperature (25°C).

Y. Okada

Papers

Physiological and biochemical changes in frog sciatic nerve during anoxia and recovery.