Isoform 3 of the Na⁺/H⁺exchanger (NHE3) is responsible for the majority of the reabsorption of NaCl, NaHCO₃, and, consequently, water in the renal proximal tubule. As such, this transporter plays an essential role in acid-base balance and extracellular fluid volume homeostasis and determining systemic arterial blood pressure levels. NHE3 activity is modulated by a number of mechanisms, including the redistribution of the transporter between the body of the microvilli (where NHE3 is active) and the base of the microvilli (where NHE3 is less active). Although the physiological, pathophysiological, and pharmacological importance of the subcellular distribution of NHE3 has been well established, the exact mechanism whereby NHE3 is translocated along microvilli microdomains of the proximal tubule apical membrane is unknown. Nonmuscle myosin IIA and unconventional myosin VI move cargoes in anterograde and retrograde directions, respectively, and are known to redistribute along with NHE3 in the proximal tubule in response to a variety of natriuretic and antinatriuretic stimuli, including stimulation or inhibition of the renin-angiotensin system, high dietary Na⁺intake, and high blood pressure. Therefore, this review aims to discuss the current evidence that suggests a potential role of myosin IIA and myosin VI in mediating the subcellular distribution of NHE3 along the kidney proximal tubule microvilli and their possible contribution in modifying NHE3-mediated Na⁺reabsorption under both physiological and pathophysiological conditions.