Recent studies have shown that the extramuscular connective tissue (ECT) is thickened and stiffened in delayed onset muscle soreness (DOMS). However, contrarily to the normal population, severe DOMS is rare in athletes or highly trained individuals. The present randomized, controlled trial therefore aimed to investigate pain as well as microcirculation and stiffness of the ECT and the erector spinae muscle following submaximal eccentric trunk extension exercise not causing DOMS. The effect of manual treatment by a therapist (myofascial release; MFR) on these parameters was to be studied. Trained healthy participants (n = 21; 31.3 ± 9.6 years; > 4 h exercise per week) performed submaximal eccentric exercise of the trunk extensors. One group was manually treated (n = 11), while the other group (n = 10) received placebo treatment with sham laser therapy. Stiffness of the ECT and the erector spinae muscle (shear wave elastography), microcirculation (white light and laser Doppler spectroscopy), palpation pain (100 mm visual analogue scale, VAS) and pressure pain threshold (indentometry, PPT) were assessed before (t₀), 24 h (t₂₄) and 48 h (t₄₈) after conditions. Erector spinae muscle stiffness increased after eccentric exercise from t₀ to t₂₄ (0.875 m/s) and from t₀ to t₄₈ (0.869 m/s). After MFR, erector spinae muscle stiffness decreased in contrast to placebo treatment at t₂₄ (-0.66 m/s), while ECT stiffness remained unchanged. Oxygen saturation increased (17-20.93%) and relative haemoglobin decreased (-9.1 - -12.76 AU) after eccentric exercise and MFR differed from placebo treatment at t₄₈ (-3.71 AU). PPT differed after MFR from placebo treatment at t₄₈ (20.69 N/mm), while VAS remained unchanged. Multiple linear regression showed that ECT stiffness and group membership predicted erector spinae muscle stiffness. MFR could have a positive effect on pain, microcirculation and muscle stiffness after submaximal eccentric exercise, suggesting better recovery, which needs to be confirmed by future work.