Three-dimensionality has been proven extensively to be critical in the development of a reliable model for different anatomical compartments and for many diseases. Currently, we can produce implantable structures that help in the regeneration of different tissues such as bone and heart. Different is the situation when we consider the neuronal compartment. As it is still difficult to understand exactly how the brain computes, to conceive how the complex chain of neuronal events can generate conscious behavior, a comprehensive and workable model of neuronal tissue still has to be found. In this perspective, in the present work, we developed and compared different 3D scaffolds to understand the effects produced by the mechanical and material properties of four different scaffolds on a 3D neuronal network. To help in preclinical testing procedure, the scalability and ease-of-use of the different approaches were also taken into consideration.Clinical Relevance- By comparing different 3D scaffolds for the creation of neuronal constructs, the results in this paper move towards understanding the best strategy to develop functional 3D neuronal units for reliable pre-clinical studies.