Exploring modulation of action potential firing by artificial graft of fast GABAergic autaptic afferences in hypophyseal neuroendocrine melanotrope cells.

Excitable cells are connected via electrical and chemical synapses to form a complex plastic network that transmit and modify action potential trains. In vivo and in vitro studies suggest that powerful type A GABAergic self-innervations, known as autapses, play a central role in the shaping of spike discharges. Herein, we have investigated the effects of artificial autaptic activity on action potential firing in cultured hypophyseal neuroendocrine melanotrope cells removed from any synaptic input. Type A autaptic conductances were introduced by using a dedicated dynamic-clamp device, based on a digital signal processor. The results indicate that cells grafted with autaptic dynamic-clamp are subjected to a modulation of both evoked firing and artificial GABA(A)-currents. The autaptic feedback reduced the action potentials amplitude, decreasing both the overshoot and the after-hyperpolarization potential (AHP). In return, the reduced voltage changes diminished the autaptic current amplitudes. The overall effect was a decrease of the cell firing rate. The introduction of a variable autaptic delay strongly altered the cell responses. Under certain conditions, the artificial autaptic current formed an additional component of the AHP which was markedly augmented. This action resulted in a stabilization of the action potential train leading to a more regular firing. In conclusion, it appeared that the autaptic feedback was very dependent on functional parameters such as the autaptic distance. Further investigations are in progress to complete our model, taking the autaptic plasticity into account.

Characterisation of Mytilus edulis hemocyte subpopulations by single cell time-lapse motility imaging.

In bivalve molluscs, defence against pathogens mainly relies on fast tissue infiltration by immunocompetent hemocytes that migrate from circulating hemolymph to sites of infection, in order to deliver, in situ, an effective immune response. In the present work, we have investigated dynamics of hemocyte subpopulations motility by combining flow cytometry coupled to Coulter-type cell volume determination, Hoffman modulation contrast microscopy, time-lapse imaging and off-line analysis of cell shape changes. Our results revealed fast modifications of hemocyte aspect in vitro, with bidirectional transitions from spread outlines to condensed cell body morphologies, in the minute range. Amoeboid or non-amoeboid types of locomotion were observed, depending on the cell shapes and on the cell subtypes, with velocities reaching up to 30 mum min(-1). Correlations between motion profiles, Hemacolor staining and flow cytometry analysis on living cells help to propose a functional mussel hemocyte classification including the motile properties of these cells. In particular, basophils were shown to be involved in dynamic hemocyte-hemocyte interactions and in the constitution of aggregation cores. Physiological implications, in terms of immune response in organisms devoid of endothelium-closed vascular system, and potential applications of hemocyte motility studies for the development and the interpretation of experiments involving hemocytes in the field of marine ecotoxicology are discussed.