A Regulatory Role for Sphingolipids in Neuronal Growth

doi:10.1074/jbc.270.18.10990

A Regulatory Role for Sphingolipids in Neuronal Growth

INHIBITION OF SPHINGOLIPID SYNTHESIS AND DEGRADATION HAVE OPPOSITE EFFECTS ON AXONAL BRANCHING (*)

From the (1) Department of Membrane Research and Biophysics, Weizmann Institute of Science, Rehovot 76100, Israel

** Incumbent of the Recanati Career Development Chair in Cancer Research. To whom correspondence should be addressed. Tel.: 972-8-342704; Fax: 972-8-344112; E mail: BMFUTER{at}WEIZMANN.WEIZMANN.AC.IL.

Abstract

Sphingolipids, particularly gangliosides, are enriched in neuronal membranes where they have been implicated as mediators of various regulatory events. We recently provided evidence that sphingolipid synthesis is necessary to maintain neuronal growth by demonstrating that in hippocampal neurons, inhibition of ceramide synthesis by Fumonisin B₁ (FB₁) disrupted axonal outgrowth (Harel, R. and Futerman, A. H. (1993) J. Biol. Chem. 268, 14476-14481). We now analyze further the relationship between neuronal growth and sphingolipid metabolism by examining the effect of an inhibitor of glucosylceramide synthesis, D-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP) and by examining the effects of both FB₁ and PDMP at various stages of neuronal development. No effects of FB₁ or PDMP were observed during the first 2 days in culture, but by day 3 axonal morphology was significantly altered, irrespective of the time of addition of the inhibitors to the cultures. Cells incubated with FB₁ or PDMP had a shorter axon plexus and less axonal branches. FB₁ appeared to cause a retraction of axonal branches between days 2 and 3, although long term incubation had no apparent effect on neuronal morphology or on the segregation of axonal or dendritic proteins. In contrast, incubation of neurons with conduritol B-epoxide, an inhibitor of glucosylceramide degradation, caused an increase in the number of axonal branches and a corresponding increase in the length of the axon plexus. A direct correlation was observed between the number of axonal branch points per cell and the extent of inhibition of either sphingolipid synthesis or degradation. These results suggest that sphingolipids play an important role in the formation or stabilization of axonal branches.

Footnotes

↵^§ Both authors contributed equally to this study.
↵^¶ Permanent address: Dept. of Biochemistry, University of Bath, Bath, Avon BA2 7AY, United Kingdom.
↵* This work was supported by the Basic Research Foundation of the Israel Academy of Science and Humanities, by Grant 91-00278 from the United States-Israel Binational Science Foundation, Jerusalem, Israel, and by the Minerva Foundation, Munich/Germany. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

SLs

sphingolipids

CBE

conduritol B-epoxide

Cer

ceramide

FB₁

Fumonisin B₁

GlcCer

glucosylceramide

GMs

gangliosides

GPI

glycosylphosphatidylinositol

[¹⁴C]hexanoyl GlcCer

N-(1-[¹⁴C]hexanoyl)-D- erythro-glucosylsphingosine

PDMP

1-phenyl-2-decanoylamino-3-morpholino-1-propanol

SM

sphingomyelin.
↵² A. Sofer and A. H. Futerman, J. Biol. Chem., in press.
↵³ I. Meivar-Levy, A. Bershadsky, and A. Futerman, unpublished observation.
↵⁴ A. Schwarz and A. H. Futerman, unpublished observations.