A new preparation for the study of cell spreading and locomotion was investigated, using time-lapse cinemicrography and photomicrography. The preparation of limpet haemocytes has the advantage of being a natural one, not derived by tissue dissociation, and is functional at normal laboratory temperatures.
The commonest cell type, the amoebocytes, have numerous spikes, up to 15 µm in length, which project from their surface, carry lamellar extensions between them, and run deep into the cytoplasm. In the spread cell this spike-supported lamella constitutes the margin of the cell, resulting in a crenate appearance. The extensions have the properties of lamellipodia, and spreading and locomotion both occur by extension, first of spikes then lamella. It is suggested that the direction of movement may be regulated by the spikes. Contact inhibition of movement is accompanied by disappearance of the spikes in the region of contact between cells and production of spikes in other regions which lead the cells away from the contact.
Spreading of cells from large aggregates of amoebocytes entails rapid centrifugal locomotion of the peripheral cells which is thought to be a consequence of contact inhibition. However, there also appears to be an opposing influence which largely prevents complete loss of contact between cells, resulting in an epithelioid morphology in a spread culture.
Amoebocytes are not adhesive to the substrate in Ca2+ and Mg2+-free (CMF) conditions but rapidly regain adhesiveness and spread normally on addition of either divalent cation to the concentration normally found in seawater. Spikes and lamellae are produced in CMF conditions but do not spread on to the substrate and do not show their normal reactions to contact with other cells.
Macrophages spread as disks of gradually increasing diameter without the aid of spikes but with the formation of conspicuous ruffle-like thickenings. Once fully spread they become quiescent and show little or no further ruffling and undergo no significant locomotion. The attachment and spreading of these cells was found to be independent of the presence in the medium of divalent cations.