Extracellular matrix glycoproteins inhibit neurite production by cultured neurons

We have analyzed the role of extracellular matrix glycoproteins in the formation of a bipolar outgrowth pattern of identified leech neurons in culture. Adult anterior pagoda (AP) neurons cultured on the inner surface of the ganglion capsules that surround central nervous system, generate two processes oriented in opposite directions. This pattern differs from those produced by these neurons cultured on other substrates, and is similar to the pattern of developing AP neurons at embryonic day 10. We used different lectins to identify subsets of glycoproteins in the extracellular matrix (ECM) of the capsules and to study their contribution to the formation of the bipolar outgrowth pattern. ECM glycoproteins binding to peanut agglutinin (PNA) or Galanthus nivalis aglutinin (GNA) lectins were detected in ganglion capsules and in ganglion extracts that had been separated by electrophoresis and blotted to nitrocellulose membranes. Four protein bands bound to PNA lectin and six other bands, including laminin subunits, bound to GNA lectin. Other lectins failed to recognize any of the proteins. For AP neurons cultured on capsules, addition of PNA lectin to the culture medium produced a dose-dependent increase in the number of primary neurites without affecting their shape, length or number of branch points. However, PNA lectin used as substrate did not affect sprouting of AP neurons. Our results suggest that PNA-binding extracellular matrix glycoproteins regulate the formation of the bipolar pattern of AP neurons by inhibiting the formation of neurites.