Neuroendocrine differentiation (NED) is a constellation of histological and functional features of certain cancer types, including prostate cancer (PCa), whereby a fraction of tumor cells display neuronal and endocrine features. Unlike patients with conventional PCa, whose tumors show a pure population of tumor cells expressing epithelial markers, patients with NED also show a subpopulation of tumor cells expressing neuroendocrine (NE) lineage markers. NED can be induced by various factors, including therapy (which further explains the association between NED and therapy resistance) and paracrine factors secreted by bone marrow stromal cell lines (which suggests that NED might be important for PCa metastasis to bone). When NED is induced in vitro, the cultured PCa cells not only show NE lineage markers but also NE lineage morphology, which is characterized by neurite-like processes. However, despite the fact that NED has been studied for several decades, the precise nature of these processes remains unclear and the term itself is thus inherently vague. This work aimed to assess the overlap between PCa NED and true neuronal phenotype. Literature search was conducted to construct a battery of pan-neuronal markers, which were assessed in NED cells in vitro by immunocytochemistry (ICC). The presence of only two core pan-neuronal markers (partly overlapping with therapy resistance) coupled with the absence of other critical pan-neuronal markers suggested that NED cells did not display a true neuronal phenotype. However, the morphology of neurite-like processes showed similarities to tunneling nanotubes (TNTs). TNTs are recently described, cytoskeletal filament-containing intercellular bridges which are involved in cell-cell trafficking of various particles ranging from cytosolic proteins to viruses and mitochondria. Time-lapse imaging, co-culture with bone marrow stromal cells, carbocyanine dye transfer, and cytoskeletal live staining were performed on NED cells in vitro, which collectively suggested that neurite-like processes mediated formation of TNT connections between NED cells. Analysis of gene expression profiles of NED was conducted using data derived from this group’s previous work as well as publicly available databases, and a set of candidate genes potentially important for TNT formation in NED cells was constructed. Most upregulated genes in the gene set were also highly expressed in brain, and several were involved in cytoskeletal organization, including neurite formation or maintenance. The expression of the corresponding proteins was assessed in NED cells in vitro by ICC, and a subset thereof was further assessed in human PCa tissue samples by immunohistochemistry-immunofluorescence (IHC-IF). Collectively, these findings suggest that NED cells do not adopt a true neuronal phenotype, but rather use subroutines of the neuronal program for extending neurite-like processes, which then mediate establishment of TNT connections.