Heteroarm core cross-linked star (CCS) polymers consist of two different polymer chains covalently joined to a cross-linked core. We investigated their self-assembly behavior to understand whether intramolecular segregation can be induced during synthesis, to produce spatial domains enriched with each polymer, and whether they would exhibit well-defined microphase separation morphologies as a result. Heteroarm CCS polymers containing polylactide (PLA) and polystyrene (PS) arms were synthesized by reversible addition-fragmentation chain transfer copolymerization of styrene and 1,2-bis(maleimidoethane) in the presence of a PLA-macro chain transfer agent (PLA-CTA), followed by chain extension with styrene (the in-out route). Dynamic light scattering, transmission electron microscopy, and small angle X-ray scattering analyses were employed to examine the self-assembly behavior in toluene and acetonitrile, as a relatively neutral and a PLA-selective solvent, respectively. Above a critical PS molar mass, lamellar-like and spherical morphologies were observed, formed by microphase separation into discrete PLA and PS domains. The increase in order with increasing PS molar mass was consistent with the segregation strength-dependent microphase separation behavior. In contrast, when the CCS polymer was synthesized by simultaneously joining PLA and PS chains (the multi macroinitiatior route) it produced rather ill-defined self-assemblies, suggesting that styrene chain extension via the in-out process is important to achieve intramolecular segregation. Using the more PLA-selective acetonitrile as a polymerization solvent indeed produced more well-defined supermicelles with PS cores and PLA coronas, confirming that intramolecular segregation can be driven by the incompatibility of the growing PS to the intramolecular environment, including PLA and the solvent.