Analysis of electrotonic coupling in patterned neuronal networks

Microcontact printing of laminin is known as an efficient approach for guiding neuronal cell migration and neurite outgrowth on artificial surfaces. In the present study, ultrathin (∼250 μm) brain stem slices of Sprague-Dawley rats (E15-E18) were cultured on laminin-patterned substrates such that neuronal cells migrating out of the slices formed grid-shaped neuronal networks along the geometry defined by the pattern. The interconnections between neighbouring pairs of neurons within these artificial networks were assessed electrophysiologically by double patch-clamp recordings and optically by microinjection of fluorescent dyes. Both functional and electrotonic synapses were detected. Based on the recorded data and simulations in PSpice, an electrical model for electrotonically coupled cells was derived. In this model the neuritic pathway is described as a cylindric cable and gap junctions are represented by an ohmic resistor. Applying this model in the data analysis the average inner radius of neurites could be determined to be ∼0.1 μm. In addition, evidence was found for a correlation between the pathwidth of the applied pattern and the diameter of neurites growing along these paths.