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Inhibition of fast axoplasmic transport by acrylamide
ENVIRONMENTAL
RESEARCH
Inhibition ROGER Laboratory
17, 251-255 (19%)
of Fast Axoplasmic L.
WEIR,
GEORGE
Transport
GLAUBIGER,
of Neuropharmacology.
NlNCDS,
AND NIH,
by Acrylamide
THOMAS Bethesda,
N. Maryland
CHASE 20014
Received July 5, 1977 The effect of acrylamide on fast axoplasmic transport in the cat sciatic nerve was studied in viva and in vitro by the technique of isotope injection. [Wlleucine was injected into the lumbar dorsal root ganglia. In cats fed daily doses of acrylamide over 1 to 4 weeks, the rate of transport was 286 mm/day compared with 424 mm/day in controls. In vitro incubation of the sciatic nerve with acrylamide (2 x IO-* M and 2 x 1O-4 M) demonstrated no such inhibition. Study of the results and review of the literature suggest that acrylamide acted subacutely on metabolism of amino acid and protein in the nerve axon, probably in the smooth endoplasmic reticulum.
INTRODUCTION
Axoplasmic transport has been identified in the peripheral nerves of several mammalian species and also in human sural nerves (Kirkpatrick and Stern, 1973; Brimijoin et al., 1973). Most of the axoplasmic flow is cellifugal from perikaryon outward along axon and dendrites providing protein, neurotransmitters, and trophic factors at the periphery of the neuron. A disturbance in this outward flow from the cell body has often been postulated as a cause for neuronal dysfunction. This has been suggested particularly for peripheral neuropathies such as occur in association with thiamine deficiency and diabetes mellitus. A decrease in rate of the slow axoplasmic transport (Pleasure et al., 1969) and the fast axoplasmic transport (Brimijoin et al., 1973; Mendell et al., 1976) have previously been correlated with a peripheral neuropathy. Acrylamide, a well-known neurotoxin, has produced a peripheral neuropathy in humans and animals. The neurotoxic effect is time- and dose-related (Kuperman, 1958)and the neuropathy occurs in the pattern of an axonal degeneration (Prineas, 1969; Suzuki and Pfaff, 1973). We describe here the effect of acrylamide on fast axoplasmic transport in the cat sciatic nerve in the intact animal and also in an incubation chamber. MATERIALS
AND METHODS
In the in vivo experiments, 75mg doses of acrylamide (Eastman Kodak) were added each day to the cats’ food over a period of 1to 4 weeks. The amount of food eaten was variable and at times diminished significantly after neurologic dysfunction developed. Cats weighed from 1.2 to 3.9 kg, and seven animals were fed acrylamide, while six others were used as controls. The technique of measuring axoplasmic transport was a modification of that described by Ochs and Hollingsworth (1971). Following pentobarbital anesthesia, a lower lumbar laminectomy was performed. [r4C]leucine (25 mCi in 10 ~1 of normal saline) was slowly injected into paired dorsal root ganglia at the L6 or L7 level. [14C]leucine of specific activity 311 mCi/mmole was obtained from Amersham. In the earlier experiments, one of 251 0013-9351/78/0172-0251$02.00/O Copyright 6 1978 by Academic Press. Inc. All rights of reproduction in any form reserved.
252
WEIR,
GLAUBIGER,
AND
CHASE
the sciatic nerves was ligatured and thus simple hematogenous spread of the isotope was characterized. Three to seven hours after radioisotope injection, the sciatic nerves were removed, blotted dry, cut into l-cm sections and weighed. Nerve sections were then solubilized by incubating with 2 ml of NCS (Amersham Searle) and 250 ~1 water at 50°C for 16 hr. The resulting solution was brought up to 20 ml with a 6% solution of spectrafluor in toluene and 5-min counts of radioactivity were made. In Vitro Three cats were anesthetized and injected with [ 14C]leucine as above. Two and one-half hours later both sciatic nerves with their ganglia were removed and incubated for up to 4.5 hr. Incubation was at 37°C in media aerated with 95% 0, and 5% CO,. One sciatic nerve (control) was simply incubated with the standard media (below). The contralateral sciatic nerve was incubated in medium to which acrylamide had been added. Concentration of acrylamide was 2 x lo-* M (14 mg/liter) or 2 x 10V2 M (1400 mg/liter). The incubation medium contained: 90.7 mM NaCl, 4.5 mM KCl, 1.15 mM KH,PO,, 2.4 mM CaCl,, 1.14 mM MgSO,*7H,O; 4.7 mM Na pyruvate, 5.2 mM fumaric acid, 4.7 mM glutamic acid, 11 mM glucose, and 23.8 mM Na bicarbonate. Six to seven hours after the ganglia were injected, the nerves were sectioned into 5-mm portions and processed as in the in vivo experiment. RESULTS
After 4 to 6 days on oral acrylamide, the cats developed head tremor, and with additional doses of acrylamide, a progressive hind limb weakness was noted. The radioactivity from the incubated sciatic nerve segments was used to calculate the distance which [14C]leucine had been transported. Data from the in vivo experiments are plotted in Fig. 1. Transport rates were estimated by a linear regression by the method of least squares. Control animals transported [14C]leucine at a rate of 424 mm/day [b = 448 (mm/day) a = -24, r = 0.81, n = 71. Acrylamide treated animals transported [14C]leucine at a rate of 286 mm/day [b = 294 (mm/day), a = -8, r = 0.80, n = 81. The transport rate was significantly slower in acrylamide treated animals (P < 0.05). There was no clear difference in the quantity of [14CJleucine transported by the two groups of animals. In Vitro The distances that [14C]leucine was transported were calculated as above and were closely similar in control and treated nerves (See Fig. 2). Transport rates (mean + SD) were 349 ? 50 mm/day (n = 3) in control specimens and 347 + 48 mm/day (n = 3) in nerves incubated with acrylamide. No acute in vitro effect of acrylamide on axonal transport is discernible. It is possible, however, that with incubation of the nerve in a higher concentration of acrylamide (greater than 2 x lop2 M) or incubation of the nerve with acrylamide for a longer period of time (over 7 hr) a slowing of axonal transport could develop. DISCUSSION In a similar in vivo experiment,
Bradley and Williams
(1973) showed the peak of
AXOPLASMIC
TRANSPORT
AND
ACRYLAMIDE
253
CM.
HOURS
FIG. 1. Rates of axoplasmic transport in control and acrylamide fed animals (in vh~). Controls: Rate = 424 mm/day; r = 0.81; b = 448 mm/day. Acrylamide: Rate = 286 mm/day; r = 0.80: 6 = 294 mm/day. (0) acrylamide treated; (0) controls.
[3H]leucine transport to be slowed by repeated acrylamide administration, but the front (advancing tip) to be unaffected. The authors discuss the data and suggest the axonal transport changes were insufficient to account for the peripheral neuropathy. However, this type of experiment is relatively coarse in terms of specific agents needed in the distal nerve or at the synapse. Thus, caution is required in interpretation of the data.
FIG. 2. Distance [‘4C]leucine was transported in vitro. Controls = 349 k 50 mm/day. Acrylamide = 347 2 48 mm/day. (a) acrylamide incubation; (ES) control incubation.
254
WEIR,
GLAUBIGER,
AND
CHASE
Both the site of action of acrylamide and the ultrastructural site where fast axonal transport occurs have been the subject of considerable study and debate. In the perikaryon of the anterior horn cell, uptake of [3H]leucine was reduced by acrylamide before an ultrastructural (or clinical) neuropathy could be documented in the perikaryon or nerve axon (Asbury et al., 1973). An earlier ultrastructural study (Prineas, 1969) was also unable to differentiate between initial structural abnormality being in axon or perikaryon. Electrophysiologic studies in early acrylamide intoxication have shown abnormalities in the distal nerve/nerve terminal region (Lowndes and Baker, 1976) and also impaired discharge from muscle stretch receptors (Sumner and Asbury, 1975). The results of both these studies may well be due to reduced function in the distal nerves secondary to deficiency of trophic or other factors normally delivered by fast axoplasmic transport. Early change in the distal nerve (compared to the more proximal nerve) has previously been shown by Schaumberg et al., (1974). Several intraaxonal structures have been considered as critical for fast axoplasmic flow. These structures include neurofilaments (Lasek, 1970), microtubules (Ochs, 1972), marginal axoplasm (Byers, 1974), and smooth endoplasmic reticulum (Byers, 1974; Colman et at., 1976; Droz ef al., 1975). Acrylamide administration has produced accumulation of neurotilaments (Prineas, 1969; Suzuki and Pfaff, 1973). More recent electron microscopic radioautography studies suggest neurofilaments have no direct role in fast axoplasmic transport though it has been suggested that accumulation of neurofilaments may obstruct fast axoplasmic transport (Mendell et al., 1976). Acrylamide administration has produced disturbance in the smooth endoplasmic reticulum (SER) (Griffin et al., 1976) and “accumulated tubulovesicular profiles” (Schaumberg et al., 1974). Present data indicate it is most likely that fast axonal transport takes place in association with the SER (Byers, 1974; Colman et al., 1976; Droz et al., 1975). The disturbance in the SER would appear to be the most identifiable cause of the slowing of fast axonal transport. The SER runs longitudinally from axon hillock to axon terminal and includes phospolipid and protein in its membranous structure. Hashimoto and Aldridge (1970) has shown that acrylamide will disturb metabolism of neural protein and the incorporation of amino acids into neural tissue (Hashimoto and Ando, 1973). Disturbance of metabolism in or near the SER probably produces the slowing of fast axonal transport, and the metabolic changes noted by Hashimoto may be the key disturbances. REFERENCES Asbury, A. K., Cox, S. C., and Kanada, D. (April 1973). “H Leucine incorporation in acrylamide neuropathy in the mouse. Presented at the annual meeting of the American Academy of Neurology. Boston. Bradley, W. G., and Williams, H. H. (1973). Axoplasmic flow in axonal neuropathies I. axoplasmic flow in cats with toxic neuropathies. Brain 96, 235-246. Brimijoin, S., Capek, P., and Dyck, P. J. (1973). Axonal transport of dopamine P-hydroxylase by human sural nerve in vitro. Science 180, 129S- 1297.
AXOPLASMIC
TRANSPORT
AND
ACRYLAMIDE
255
Byers, M. (1974). Structural correlates of rapid axonal transport: evidence that microtubules may not be directly involved. Brain Ras. 75, 97- 113. Colman, D. R., Scalia, F., and Cabrales, E. (1976). Light and electron microscopic observation on the anterograde transport of horseradish peroxidase in the optic pathway in the mouse and rat. Brain Res. 102, 156-163. Droz, B., Rambourg, A., and Koenig, H. L. (1975). The smooth endoplasmic reticulum; structure and role in the renewal of axonal membrane and synaptic vesicles by fast axonal transport. Brain Res. 93, I-13. Griffin, J. W., Price, D. L., and Drachman, D. B. (April 1976). Impaired regeneration in acrylamide neuropathy: Role of axonal transport. Presented at the annual meeting of the American Academy of Neurology. Toronto, Canada. Hashimoto, K., and Aldridge, W. M. (1970). Biochemical studies on acrylamide, a neurotoxic agent. Biochem. Pharmacol. 19, 2591-2604. Hashimoto, K., and Ando, K. (1973). Alteration of amino acid incorporation into proteins of nervous system in vitro after administration of acrylamide to rats. Biochem. Pharmacol. 22, 1057- 1066. Kaplan, M., Murphy, S. D., and Gilles, F. H. (1973). Modification of acrylamide neuropathy in rats by selected factors. Toxicol. Appl. Ph&macol. 24, X4-570. Kirkpatrick, J. B., and Stern, L. Z. (1973). Axoplasmic flow in human sural nerve. Arch. Neural. 28, 308-3 12. Kuperman, A. S. (1958). Effects of acrylamide on the CNS of the cat. J. Pharmacol. Exp. Thu. 123, 180- 192. Lasek, R. J. (1970). Protein transport in neurons. Inr. Rev. Neurobiol. 13, 289-324. Lowndes, H. E., and Baker, T. (1976). Studies on drug-induced neuropathies: III, motor nerve deficit in cats with experimental acrylamide neuropathy. Eur. J. Pharmacol. 35, 177-184. Mendell, J. R., Saida, K., Weiss, H. S., and Savage, R. (April 1976). Methyl n-butyl ketone-induced changes in fast axoplasmic transport. Presented at the annual meeting of the American Academy of Neurology. Toronto, Canada. Ochs, S., and Hollingsworth, D. (1971) Dependence of fast axoplasmic transport in nerve on oxidative metabolism. J. Neurochem. 18, 107- 114. Ochs, S. (1972) Fast transport of materials in mammalian nerve fibres. Science 176, 252-260. Pleasure, D. E., Mishler, K. C., and King-Engel, W. (1969). Axonal transport of proteins in experimental neuropathies. Science 166, 524-525. Prineas, J. ( 1969). The pathogenesis of dying-back polyneuropathies. II. An ultrastructural study of experimental acrylamide intoxication in the cat. J. Neuroparhol. Exp. Neural. 28, 598-621. Schaumberg, H. H., Wisniewski, H., and Spencer, P. S. (April 1974). An ultrastructural study of the dying-back process. Presented at the annual meeting of the American Academy of Neurology. San Francisco. Sumner, A. J., and Asbury, A. K. (1973). Physiological studies of the dying-back phenomenon: muscle stretch afferents in acrylamide neuropathy. Brain 98, 91- 100. Suzuki, K., and Pfaff, L. D. (1973). Acrylamide neuropathy in rats. An electron microscopic study of degeneration and regeneration. Acra Neuropathol (Berlin), 24, 197-213.
RESEARCH
Inhibition ROGER Laboratory
17, 251-255 (19%)
of Fast Axoplasmic L.
WEIR,
GEORGE
Transport
GLAUBIGER,
of Neuropharmacology.
NlNCDS,
AND NIH,
by Acrylamide
THOMAS Bethesda,
N. Maryland
CHASE 20014
Received July 5, 1977 The effect of acrylamide on fast axoplasmic transport in the cat sciatic nerve was studied in viva and in vitro by the technique of isotope injection. [Wlleucine was injected into the lumbar dorsal root ganglia. In cats fed daily doses of acrylamide over 1 to 4 weeks, the rate of transport was 286 mm/day compared with 424 mm/day in controls. In vitro incubation of the sciatic nerve with acrylamide (2 x IO-* M and 2 x 1O-4 M) demonstrated no such inhibition. Study of the results and review of the literature suggest that acrylamide acted subacutely on metabolism of amino acid and protein in the nerve axon, probably in the smooth endoplasmic reticulum.
INTRODUCTION
Axoplasmic transport has been identified in the peripheral nerves of several mammalian species and also in human sural nerves (Kirkpatrick and Stern, 1973; Brimijoin et al., 1973). Most of the axoplasmic flow is cellifugal from perikaryon outward along axon and dendrites providing protein, neurotransmitters, and trophic factors at the periphery of the neuron. A disturbance in this outward flow from the cell body has often been postulated as a cause for neuronal dysfunction. This has been suggested particularly for peripheral neuropathies such as occur in association with thiamine deficiency and diabetes mellitus. A decrease in rate of the slow axoplasmic transport (Pleasure et al., 1969) and the fast axoplasmic transport (Brimijoin et al., 1973; Mendell et al., 1976) have previously been correlated with a peripheral neuropathy. Acrylamide, a well-known neurotoxin, has produced a peripheral neuropathy in humans and animals. The neurotoxic effect is time- and dose-related (Kuperman, 1958)and the neuropathy occurs in the pattern of an axonal degeneration (Prineas, 1969; Suzuki and Pfaff, 1973). We describe here the effect of acrylamide on fast axoplasmic transport in the cat sciatic nerve in the intact animal and also in an incubation chamber. MATERIALS
AND METHODS
In the in vivo experiments, 75mg doses of acrylamide (Eastman Kodak) were added each day to the cats’ food over a period of 1to 4 weeks. The amount of food eaten was variable and at times diminished significantly after neurologic dysfunction developed. Cats weighed from 1.2 to 3.9 kg, and seven animals were fed acrylamide, while six others were used as controls. The technique of measuring axoplasmic transport was a modification of that described by Ochs and Hollingsworth (1971). Following pentobarbital anesthesia, a lower lumbar laminectomy was performed. [r4C]leucine (25 mCi in 10 ~1 of normal saline) was slowly injected into paired dorsal root ganglia at the L6 or L7 level. [14C]leucine of specific activity 311 mCi/mmole was obtained from Amersham. In the earlier experiments, one of 251 0013-9351/78/0172-0251$02.00/O Copyright 6 1978 by Academic Press. Inc. All rights of reproduction in any form reserved.
252
WEIR,
GLAUBIGER,
AND
CHASE
the sciatic nerves was ligatured and thus simple hematogenous spread of the isotope was characterized. Three to seven hours after radioisotope injection, the sciatic nerves were removed, blotted dry, cut into l-cm sections and weighed. Nerve sections were then solubilized by incubating with 2 ml of NCS (Amersham Searle) and 250 ~1 water at 50°C for 16 hr. The resulting solution was brought up to 20 ml with a 6% solution of spectrafluor in toluene and 5-min counts of radioactivity were made. In Vitro Three cats were anesthetized and injected with [ 14C]leucine as above. Two and one-half hours later both sciatic nerves with their ganglia were removed and incubated for up to 4.5 hr. Incubation was at 37°C in media aerated with 95% 0, and 5% CO,. One sciatic nerve (control) was simply incubated with the standard media (below). The contralateral sciatic nerve was incubated in medium to which acrylamide had been added. Concentration of acrylamide was 2 x lo-* M (14 mg/liter) or 2 x 10V2 M (1400 mg/liter). The incubation medium contained: 90.7 mM NaCl, 4.5 mM KCl, 1.15 mM KH,PO,, 2.4 mM CaCl,, 1.14 mM MgSO,*7H,O; 4.7 mM Na pyruvate, 5.2 mM fumaric acid, 4.7 mM glutamic acid, 11 mM glucose, and 23.8 mM Na bicarbonate. Six to seven hours after the ganglia were injected, the nerves were sectioned into 5-mm portions and processed as in the in vivo experiment. RESULTS
After 4 to 6 days on oral acrylamide, the cats developed head tremor, and with additional doses of acrylamide, a progressive hind limb weakness was noted. The radioactivity from the incubated sciatic nerve segments was used to calculate the distance which [14C]leucine had been transported. Data from the in vivo experiments are plotted in Fig. 1. Transport rates were estimated by a linear regression by the method of least squares. Control animals transported [14C]leucine at a rate of 424 mm/day [b = 448 (mm/day) a = -24, r = 0.81, n = 71. Acrylamide treated animals transported [14C]leucine at a rate of 286 mm/day [b = 294 (mm/day), a = -8, r = 0.80, n = 81. The transport rate was significantly slower in acrylamide treated animals (P < 0.05). There was no clear difference in the quantity of [14CJleucine transported by the two groups of animals. In Vitro The distances that [14C]leucine was transported were calculated as above and were closely similar in control and treated nerves (See Fig. 2). Transport rates (mean + SD) were 349 ? 50 mm/day (n = 3) in control specimens and 347 + 48 mm/day (n = 3) in nerves incubated with acrylamide. No acute in vitro effect of acrylamide on axonal transport is discernible. It is possible, however, that with incubation of the nerve in a higher concentration of acrylamide (greater than 2 x lop2 M) or incubation of the nerve with acrylamide for a longer period of time (over 7 hr) a slowing of axonal transport could develop. DISCUSSION In a similar in vivo experiment,
Bradley and Williams
(1973) showed the peak of
AXOPLASMIC
TRANSPORT
AND
ACRYLAMIDE
253
CM.
HOURS
FIG. 1. Rates of axoplasmic transport in control and acrylamide fed animals (in vh~). Controls: Rate = 424 mm/day; r = 0.81; b = 448 mm/day. Acrylamide: Rate = 286 mm/day; r = 0.80: 6 = 294 mm/day. (0) acrylamide treated; (0) controls.
[3H]leucine transport to be slowed by repeated acrylamide administration, but the front (advancing tip) to be unaffected. The authors discuss the data and suggest the axonal transport changes were insufficient to account for the peripheral neuropathy. However, this type of experiment is relatively coarse in terms of specific agents needed in the distal nerve or at the synapse. Thus, caution is required in interpretation of the data.
FIG. 2. Distance [‘4C]leucine was transported in vitro. Controls = 349 k 50 mm/day. Acrylamide = 347 2 48 mm/day. (a) acrylamide incubation; (ES) control incubation.
254
WEIR,
GLAUBIGER,
AND
CHASE
Both the site of action of acrylamide and the ultrastructural site where fast axonal transport occurs have been the subject of considerable study and debate. In the perikaryon of the anterior horn cell, uptake of [3H]leucine was reduced by acrylamide before an ultrastructural (or clinical) neuropathy could be documented in the perikaryon or nerve axon (Asbury et al., 1973). An earlier ultrastructural study (Prineas, 1969) was also unable to differentiate between initial structural abnormality being in axon or perikaryon. Electrophysiologic studies in early acrylamide intoxication have shown abnormalities in the distal nerve/nerve terminal region (Lowndes and Baker, 1976) and also impaired discharge from muscle stretch receptors (Sumner and Asbury, 1975). The results of both these studies may well be due to reduced function in the distal nerves secondary to deficiency of trophic or other factors normally delivered by fast axoplasmic transport. Early change in the distal nerve (compared to the more proximal nerve) has previously been shown by Schaumberg et al., (1974). Several intraaxonal structures have been considered as critical for fast axoplasmic flow. These structures include neurofilaments (Lasek, 1970), microtubules (Ochs, 1972), marginal axoplasm (Byers, 1974), and smooth endoplasmic reticulum (Byers, 1974; Colman et at., 1976; Droz ef al., 1975). Acrylamide administration has produced accumulation of neurotilaments (Prineas, 1969; Suzuki and Pfaff, 1973). More recent electron microscopic radioautography studies suggest neurofilaments have no direct role in fast axoplasmic transport though it has been suggested that accumulation of neurofilaments may obstruct fast axoplasmic transport (Mendell et al., 1976). Acrylamide administration has produced disturbance in the smooth endoplasmic reticulum (SER) (Griffin et al., 1976) and “accumulated tubulovesicular profiles” (Schaumberg et al., 1974). Present data indicate it is most likely that fast axonal transport takes place in association with the SER (Byers, 1974; Colman et al., 1976; Droz et al., 1975). The disturbance in the SER would appear to be the most identifiable cause of the slowing of fast axonal transport. The SER runs longitudinally from axon hillock to axon terminal and includes phospolipid and protein in its membranous structure. Hashimoto and Aldridge (1970) has shown that acrylamide will disturb metabolism of neural protein and the incorporation of amino acids into neural tissue (Hashimoto and Ando, 1973). Disturbance of metabolism in or near the SER probably produces the slowing of fast axonal transport, and the metabolic changes noted by Hashimoto may be the key disturbances. REFERENCES Asbury, A. K., Cox, S. C., and Kanada, D. (April 1973). “H Leucine incorporation in acrylamide neuropathy in the mouse. Presented at the annual meeting of the American Academy of Neurology. Boston. Bradley, W. G., and Williams, H. H. (1973). Axoplasmic flow in axonal neuropathies I. axoplasmic flow in cats with toxic neuropathies. Brain 96, 235-246. Brimijoin, S., Capek, P., and Dyck, P. J. (1973). Axonal transport of dopamine P-hydroxylase by human sural nerve in vitro. Science 180, 129S- 1297.
AXOPLASMIC
TRANSPORT
AND
ACRYLAMIDE
255
Byers, M. (1974). Structural correlates of rapid axonal transport: evidence that microtubules may not be directly involved. Brain Ras. 75, 97- 113. Colman, D. R., Scalia, F., and Cabrales, E. (1976). Light and electron microscopic observation on the anterograde transport of horseradish peroxidase in the optic pathway in the mouse and rat. Brain Res. 102, 156-163. Droz, B., Rambourg, A., and Koenig, H. L. (1975). The smooth endoplasmic reticulum; structure and role in the renewal of axonal membrane and synaptic vesicles by fast axonal transport. Brain Res. 93, I-13. Griffin, J. W., Price, D. L., and Drachman, D. B. (April 1976). Impaired regeneration in acrylamide neuropathy: Role of axonal transport. Presented at the annual meeting of the American Academy of Neurology. Toronto, Canada. Hashimoto, K., and Aldridge, W. M. (1970). Biochemical studies on acrylamide, a neurotoxic agent. Biochem. Pharmacol. 19, 2591-2604. Hashimoto, K., and Ando, K. (1973). Alteration of amino acid incorporation into proteins of nervous system in vitro after administration of acrylamide to rats. Biochem. Pharmacol. 22, 1057- 1066. Kaplan, M., Murphy, S. D., and Gilles, F. H. (1973). Modification of acrylamide neuropathy in rats by selected factors. Toxicol. Appl. Ph&macol. 24, X4-570. Kirkpatrick, J. B., and Stern, L. Z. (1973). Axoplasmic flow in human sural nerve. Arch. Neural. 28, 308-3 12. Kuperman, A. S. (1958). Effects of acrylamide on the CNS of the cat. J. Pharmacol. Exp. Thu. 123, 180- 192. Lasek, R. J. (1970). Protein transport in neurons. Inr. Rev. Neurobiol. 13, 289-324. Lowndes, H. E., and Baker, T. (1976). Studies on drug-induced neuropathies: III, motor nerve deficit in cats with experimental acrylamide neuropathy. Eur. J. Pharmacol. 35, 177-184. Mendell, J. R., Saida, K., Weiss, H. S., and Savage, R. (April 1976). Methyl n-butyl ketone-induced changes in fast axoplasmic transport. Presented at the annual meeting of the American Academy of Neurology. Toronto, Canada. Ochs, S., and Hollingsworth, D. (1971) Dependence of fast axoplasmic transport in nerve on oxidative metabolism. J. Neurochem. 18, 107- 114. Ochs, S. (1972) Fast transport of materials in mammalian nerve fibres. Science 176, 252-260. Pleasure, D. E., Mishler, K. C., and King-Engel, W. (1969). Axonal transport of proteins in experimental neuropathies. Science 166, 524-525. Prineas, J. ( 1969). The pathogenesis of dying-back polyneuropathies. II. An ultrastructural study of experimental acrylamide intoxication in the cat. J. Neuroparhol. Exp. Neural. 28, 598-621. Schaumberg, H. H., Wisniewski, H., and Spencer, P. S. (April 1974). An ultrastructural study of the dying-back process. Presented at the annual meeting of the American Academy of Neurology. San Francisco. Sumner, A. J., and Asbury, A. K. (1973). Physiological studies of the dying-back phenomenon: muscle stretch afferents in acrylamide neuropathy. Brain 98, 91- 100. Suzuki, K., and Pfaff, L. D. (1973). Acrylamide neuropathy in rats. An electron microscopic study of degeneration and regeneration. Acra Neuropathol (Berlin), 24, 197-213.