Within the first day after plating one of the short processes of a hippocampal neuron becomes defined as the single axon of the cell that grows out very rapidly. Both rate and direction of outgrowth are influenced by the nature of the substrate the axon encounters. This movie follows the elongation of the axon on a uniform substrate over a period of 16 hours. Note how the axon elongates in spurts.

Time lapse: 16.5 hours (one loop of movie)

Nanotechnology can be used to create patterned non-uniform substrates for neurons to grow on. Here we followed axon elongation on a pattern (visualized in fluorescent green) where the axon encounters a 3-way intersection. The axon makes a sharp left turn without hesitation and after a short exploratory phase extends a branch also in the original growth direction. Finally, several hours later, the cell decides to branch in the 3rd direction, again following what appears to be an exploratory phase where it senses the track.

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Time lapse: 18 hours (one loop of movie)

Functional differences of axons and dendrites hinge upon differences in their molecular composition. Membrane proteins destined to either of the domains leave the Golgi in tubulovesicular carriers that are transported by molecular motors along microtubular tracks. Currently, we are investigating the identity and transport behavior of carriers containing axonal and dendritic cell surface proteins by tagging them with fluorescent proteins (FP’s).

This neuron expresses an FP-tagged protein (NgCAM) that turns up exclusively on the axonal cell surface. Interestingly, carriers containing this axonal protein enter freely into all the processes radiating from the cell body, i.e. the axon (top right) and all dendrites. Transport of carriers is bi-directional, both away and towards the cell body. The bright spot in the cell body corresponds to the Golgi area where the carriers originate.

Time lapse: 30 seconds (one loop of movie)

This neuron expresses an FP-tagged protein (TfR) that turns up exclusively on the dendritic cell surface. Carriers for this dendritic protein show preferential transport into dendrites (all processes left of cell body shown in 1st movie; lower branch of process shown in 2nd movie) within a day of the axon growing out (upper branch in 2nd movie).

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Time lapse: 30 seconds (one loop of movie)

Later in development, when dendrites mature, transport of carriers for dendritic cell surface proteins becomes highly selective. Dendritic carriers, pseudo-colored in green, are seen traveling bi-directionally in the dendritic branch, identified retrospectively by the presence of a dendritic marker, MAP2, pseudo-colored in red. No carriers enter the axon.

Time lapse: 30 seconds (one loop of movie)

Delivery of membrane proteins to the cell surface occurs by fusion of the carriers with the plasma membrane. Two fusion events have been captured fortuitously in this movie. Upon fusion, the contents of the carriers very rapidly diffuse in the plasma membrane, resulting in a fleeting impression of a railroad track due to the optical sectioning power of the microscope.

Time lapse: 30 seconds (one loop of movie)