Prosocial helping behavior, characterized by voluntary actions taken to benefit others, plays a vital role in promoting cooperation and maintaining social bonds across human and animal social groups. In this review, we examine key conceptual issues surrounding prosocial behavior, focusing specifically on targeted helping and comforting actions. We outline the behavioral paradigms used to study these two types of prosocial behaviors and summarize recent insights into their underlying neural mechanisms. Drawing on findings across species, with an emphasis on rodent models, we discuss how these behaviors are regulated by molecularly and anatomically defined neural systems and how distinct neuronal populations and circuits may differentially regulate targeted helping and comforting behaviors. Lastly, we discuss the clinical relevance of this research by addressing the implications of prosocial deficits in psychiatric disorders.