Intramuscular injection of hypertonic saline: in vitro and in vivo muscle tissue toxicity and spinal neurone c-fos expression

Abstract

Intramuscular injection of hypertonic saline (4-6% NaCl) is widely used to induce muscle pain in volunteers. The quality of the pain is comparable to clinical muscle pain with localised and referred pain. The objective was to evaluate the muscle toxicity of hypertonic saline by characterisation of 1) cytotoxicity in vitro, 2) local muscle toxicity in rabbits and 3) number of spinal dorsal horn neurones expressing c-fos after intramuscular injection in pigs as an indicator of nociception. Rat myocyte cultures and erythrocyte suspensions were treated with hypertonic NaCl solutions. The creatine kinase activity remaining in the myocytes and haemolysis were measured. Groups of six rabbits were given an intramuscular injection of 0.5 ml of 0.9, 3 or 6% NaCl. Three days later, creatine kinase activity was determined in injection site muscle tissue and normal contralateral muscle. The amount of injection site muscle tissue totally depleted of creatine kinase was calculated. Groups of two pigs were given an intramuscular injection of 3.0 ml of 6% NaCl. The spinal cord was sampled 1, 2 or 3 hr later and processed for stereological quantification of the number of dorsal horn neurones expressing c-fos. Saline was not toxic in vitro at 0.9-6%, but toxic to erythrocytes at 7% or higher and rat myocytes at 15% or higher. No muscle toxicity was seen in rabbits. The number of dorsal horn neurones expressing c-fos was not above basal level. In conclusion, 6% saline caused no in vitro or in vivo toxicity in sensitive models. Consequently, the pain caused by intramuscular injection of hypertonic saline is most likely not related to tissue damage. Consistently, intramuscular injection of 6% NaCl did not activate dorsal horn neurones in pigs to express c-fos beyond basal level.