Research objectives and content
Recent experiments have established the presence of spontaneous oscillations in electrical activity within neuronal networks of adult and developing brain centers. This type of activity has been recorded in different structures of the nervous system, including retina, spinal cord, brainstem, thalamus and cortex. Some findings suggest that this spontaneous activity may have a critical influence upon the correct development of neuronal connections and functional properties of neuronal circuits. The principal objective of this proposal is to investigate the cellular mechanisms underlying the generation of spontaneous electrical activity in the developing brain. The chosen preparation is the isolated brainstem (and slices) of the chick embryo, at different developmental stages. Recent findings have provided clear evidence indicating repetition of neuronal networks, having similar (but not identical) rhythmogenic properties, along the rostrocaudal axis of the developing chick hindbrain. The applicant proposes to use in-vitro recording techniques, combined with Ca2+ imaging techniques and neuroanatomical tracing techniques to identify and characterise the cellular elements involved in the generation of spontaneous oscillatory motor activities.
The working hypothesis is that oscillatory motor activity depends on the activity of synaptically connected pacemaker neurons. The proposal outlines experimental strategies (extracellular recordings, Ca2+-imaging and labelling methods) to identify the location of putative pacemaker neurons within the brainstem. Following the location of pacemaker neurons, cross-correlation and spike triggered-averaging techniques will be used to establish the pattern of connectivity between pacemaker cells and other brainstem neurons, including motoneurons. In parallel experiments the applicant will investigate the intrinsic ionic mechanisms of pacemaker neurons that may contribute to their spontaneous activity. Using pharmacological techniques, the role of gabaergic and glycinergic synapses in the spread and synchronisation of spontaneous motor activity will be investigated as well. Finally, manipulation of the normal segmentation pattern of the brainstem will be used to investigate the potential role of diffusible agents in establishing the rostrocaudal gradient of spontaneous activity within the brainstem.
Training content (objective, benefit and expected impact)
This application is submitted in the context of a return (class R) fellowship. The work will be carried out at the Institute de Neurosciences (Director: Prof. Carlos Belmonte) in the University of Alicante, a recognised centre for neurobiology research in Spain. The applicant should benefit from the suggested proposal in a number of ways. It will allow training in different experimental techniques (tissue grafting, embryology) that are complementary to the ones currently used by the applicant. It will also allow him to return to his country of origin and establish collaborative work with other Spanish scientists in an important research area (developmental neuroscience). The expected results should help advance our understanding of the basic cellular mechanisms operating during vertebrate brain development, in particular the development of motor activity and motor circuits.