The technique for gene transfer and expression via retrograde axonal transport of the viral vectors enables us to introduce the transgene into neuronal populations that innervate the brain regions where the vectors are injected. Our group recently developed human immunodeficiency virus type 1-based vector for neuron-specific retrograde gene transfer (NeuRet) with pseudotyping of fusion glycoprotein type C (FuG-C), which consists of the N-terminal region (439 amino acids) of the extracellular domain of rabies virus glycoprotein (RVG) and the membrane-proximal region (16 amino acids) of the extracellular domain and the transmembrane/cytoplasmic domains of vesicular stomatitis virus glycoprotein (VSVG) (Kato et al., 2011). Although fusion in the membrane-proximal region of viral envelope glycoproteins improves the efficiency of retrograde gene transfer, the part of the junction between RVG/VSVG segments in fusion glycoproteins that confers the most efficacious gene transfer has not yet been determined. To optimize the junction of RVG and VSVG segments in fusion glycoproteins in their membrane-proximal region, we constructed various types of fusion glycoproteins and generated lentiviral vectors pseudotyped with these fusion glycoproteins. We then tested the efficiency of the pseudotyped vectors for the in vivo gene transfer through retrograde transport, comparing that of the NeuRet vector with FuG-C. We found a novel type of fusion glycoprotein, termed type E (FuG-E), that showed improved efficiency of retrograde gene transfer while retaining the property of neuron-specific transduction (Kato et al., 2014). This NeuRet vector with FuG-E will provide a powerful tool for genetic treatment of neurological and neurodegenerative diseases and for the study of neural circuit mechanisms underlying various brain functions.