Neuro-endocrine biomarkers of pollution-induced stress in marine invertebrates

Nitric oxide (NO) is a molecular mediator of the non-specific immune response of vertebrates but production has also been recorded in response to bacterial infection in invertebrates. Together with biogenic amines and cytokines, production of NO in invertebrate phagocytic haemocytes is thought to represent a putative stress response. In this study the hypothesis that NO might be a sensitive biomarker of stress in invertebrates has been tested. The spectrophotometric Griess assay for nitrite and nitrate has been modified so that the final breakdown products of NO are suitable for measurement in invertebrate haemolymph. The assay was linear in the range 1-100 microM with a sensitivity of 4 microM. The baseline NOx measurement in unstressed mussels was 0.74 mM NOx/mg protein. Mytilus edulis were kept at 15 micro C in filtered seawater for at least 2 days prior to exposure to TBTO (0.001-0.1 mg/l) for 24 h. Preliminary results suggest that TBTO perturbs the NO response and that the outlined assay protocol is a sensitive means of detecting those changes. It is proposed that NO measurements potentially offer a highly sensitive, non-invasive means of monitoring stress responses associated with environmental change.