On the Application of Cultured Neuronal Cell Lines in Neurotoxicological Studies: Implications of Acrylamide-induced Neurite Disintegration

Abstract

Summary

Cultures of the mouse neuroblastoma cell line C1300, clone N1E115 were exposed to acrylamide at 3.5 x 10^-4M for 14 days (subacute situation) or at 2.8 x 10^-3 M for 24 hr (acute situation).

In the subacute situation the total uptake of 2-deoxy-D-glucose was stimulated. This could be explained by an increase in both the non-specific diffusion and the specific transport. The activity of the glycolytic enzyme, enolase (EC4.2.1.11), was unaffected by exposure to acrylamide, whereas the activity of glyceraldehyde-3-phosphate-dehydrogenase (EC1.2.1.12) was inhibited. Acrylamide had a marked stimulating effect on the respiratory activity of the cells, whereas the incorporation of tritiated leucine remained unchanged. Furthermore, membrane integrity was maintained throughout the acrylamide exposure as judged by an unchanged rate of 2-deoxy-D-glucose-6-phosphate efflux. Corresponding results were obtained in the acute situation. In N1E115 cultures and under the experimental conditions used in this work acrylamide caused neurite degeneration resembling distal axonopathy in vivo. It is suggested that these degenerative changes are not due to a general intoxication of the cells, but rather to a specific effect. Consequently, the N1E115 cell line might be useful in studies of chemically-induced axonopathies.

Keywords

neuroblastoma cells acrylamide glucose uptake glucose metabolism leucine incorporation membrane integrity respiratory activity

Get full access to this article

View all access options for this article.

References

Walum

(1984). On the application of cultured neuronal cell lines in neurotoxicological studies: cytotoxicity of acrylamide. ATLA 11, 135–145.

Cavanagh

J.B.

(1982). The pathokinetics of acrylamide intoxication: a reassessment of the problem. Neuropathology and Applied Neurobiology 8, 315–336.

Cavanagh

J.B.

(1979). The dying back process. A common denominator in many naturally occurring and toxic neuropathies. Archive for Pathology and Laboratory Medicine 103, 659–664.

Walum

(1982). Membrane lesions in cultured mouse neuroblastoma cells exposed to metal compounds. Toxicology 25, 67–74.

Sokoloff

, Reivich

, Kennedy

, Des Rosiers

M.H.

, Patlak

C.S.

, Pettigrew

K.D.

, Sakurada

, and Shinohara

(1977). The ¹⁴C deoxyglucose method for the measurement of local cerebral glucose utilization: theory, procedure and normal values in the conscious and anaesthetised albino rat. Journal of Neurochemistry 28, 897–916.

Walum

(1975). Glucose uptake into cultured tumor cells from the nervous system. Thesis, University of Gothenburg

Sweden.

Kanje

, Walum

, & Edström

(1979). Effects of dibutyrylic cyclic AMP and cytochalasin B on cultured human glioma’ cells. Zeitschrift für Mikroskopische und Anatomische Forschung, Leipzig 93, 487–496.

Vas

, and Blatke

(1981). Evidence for absence of an interaction between purified 3-phosphoglycerate kinase and glyceraldehyde-3-phosphate dehydrogenase. Biochimica et Biophysica Acta 660, 193–198.

Rider

C.C.

, and Taylor

C.B.

(1974). Enolase isoenzymes in rat tissues. Electrophoretic, chromatographic, immunological and kinetic properties. Biochimica et Biophysica Acta 365, 285–300.

10.

Peterson

, and Walum

(1983). Growth and morphology of neuronal cell lines cultured in perfusion. In Vitro 19, 875–879.

11.

Read

S.M.

, and Northcote

D.H.

(1981). Minimisation of variation in response to different proteins of the Comassie Blue G dyebinding assay for protein. Analytical Biochemistry 116, 53–64.

12.

Fullerton

P.M.

, and Barnes

J.M.

(1966). Peripheral neuropathy in rats, produced by acrylamide. British Journal of Industrial Medicine 23, 210–221.

13.

Hashimoto

, and Aldridge

W.N.

(1970). Biochemical studies on acrylamide, a neurotoxic agent. Biochemical Pharmacology 19, 2591–2604.

14.

Walum

, & Edström

(1976). Kinetics of 2-deoxy-D-glucose transport into cultured mouse neuroblastoma cells. Experimental Cell Research 97, 15–22.

15.

Haffne

S.C.

, and Seeds

N.W.

(1975). Neuroblastoma: the E. coli of neurobiology? Life Sciences 16, 1649–1657.

16.

Sokoloff

(1981). Circulation and Energy Metabolism of the Brain. In Basic Neurochemistry (ed. Siegel

G.J.

, Alberts

R.W.

, Agranoff

B.W.

, Katzman

), third edition, pp. 471–495. Boston: Little, Brown and Company.

17.

Sabri

M.J.

, and Spencer

P.S.

(1980). Toxic Distal Axonopathy: Biochemical Studies and Hypothetical Mechanisms. In Experimental and Clinical Neurotoxicology (ed. Spencer

P.S.

, Schaumberg

H.H.

), pp. 206–219. Baltimore: Waverly Press.

18.

Walum

, and Peterson

(1982). Tritiated 2-deoxy-D-glucose as a probe for cell membrane permeability studies. Analytical Biochemistry 120, 8–11.

19.

Walum

, and Marchner

(1983). Effects of mercuric chloride on the membrane integrity of cultured cell lines. Toxicology Letters 18, 89–95.

20.

Schubert

, Harris

A.J.

, Heinemann

, Kidokosa

, Patrich

, and Steinbach

J.H.

(1973). Differentiation and Interaction of Clonal Lines of Nerve and Muscle. In Tissue Culture of the Nervous System (ed. Sato

G.-H.

), pp. 55–86. New York: Plenum Press.

21.

Keller

, Lange

, and Noske

(1981). D-glucose transport in cultured cells of neural origin: the membrane as possible control point of glucose utilisation. Journal of Neurochemistry 36, 1012–1017.

22.

Spencer

P.S.

, Mohammed

, Schaumberg

H.H.

, and Moore

Ch.

(1979). Does a defect of energy metabolism in the nerve fiber underlie axonal degeneration in polyneuropathies? Annals of Neurology 5, 501–507.

23.

Howland

R.D.

, Vyas

I.L.

, and Lowndes

H.E.

(1980a). The etiology of acrylamide neuropathy: possible involvement of neuron specific enolase. Brain Research 190, 529–535.

24.

Howland

R.D.

, Vyas

I.L.

, Lowndes

H.E.

, and Argentieri

T.M.

(1980b). The etiology of toxic peripheral neuropathies: in vitro effects of acrylamide and 2.5-hexadione on brain enolase and other glycolytic enzymes. Brain Research 202, 131–142.

25.

Marangos

P.J.

, Goodwin

F.D.

, Parma

A.M.

, Lanter

, and Trams

(1978). Neuron specific protein (NSP) in neuroblastoma cells: relation to differentiation. Brain Research 145, 49–58.