Soft alginate hydrogels support robust neurite outgrowth, but their rapid disintegration in solutions of high ionic strength restricts them from long-term in vivo applications. Aiming to enhance the mechanical stability of soft alginate hydrogels, we investigated how changes in pH and ionic strength during gelation influence the swelling, stiffness, and disintegration of a three-dimensional (3D) alginate matrix and its ability to support neurite outgrowth. Hydrogels were generated from dry alginate layers through ionic crosslinks with Ca(2+) (≤ 10 mM) in solutions of low or high ionic strength and at pH 5.5 or 7.4. High- and low-viscosity alginates with different molecular compositions demonstrated pH and ionic strength-independent increases in hydrogel volume with decreases in Ca(2+) concentrations from 10 to 2 mM. Only soft hydrogels that were synthesized in the presence of 150 mM of NaCl (Ca-alginate NaCl) displayed long-term volume stability in buffered physiological saline, whereas analogous hydrogels generated in NaCl-free conditions (Ca-alginate) collapsed. The stiffnesses of Ca-alginate NaCl hydrogels elevated from 0.01 to 19 kPa as the Ca(2+)-concentration was raised from 2 to 10 mM; however, only Ca-alginate NaCl hydrogels with an elastic modulus ≤ 1.5 kPa that were generated with ≤ 4 mM of Ca(2+) supported robust neurite outgrowth in primary neuronal cultures. In conclusion, soft Ca-alginate NaCl hydrogels combine mechanical stability in solutions of high ionic strength with the ability to support neural growth and could be useful as 3D implants for neural regeneration in vivo.