Coordination of cytoskeletal dynamics and cell behaviour during Drosophila abdominal morphogenesis.

During morphogenesis, cells exhibit various behaviours, such as migration and constriction, which need to be coordinated. How this is achieved remains elusive. During morphogenesis of the Drosophila adult abdominal epidermis, larval epithelial cells (LECs) migrate directedly before constricting apically and undergoing apoptosis. Here, we study the mechanisms underlying the transition from migration to constriction. We show that LECs possess a pulsatile apical actomyosin network, and that a change in network polarity correlates with behavioural change. Exploring the properties of the contractile network, we find that cell contractility, as determined by myosin activity, has an impact on the behaviour of the network, as well as on cytoskeletal architecture and cell behaviour. Pulsed contractions occur only in cells with intermediate levels of contractility. Furthermore, increasing levels of the small Rho GTPase Rho1 disrupts pulsing, leading to cells that cycle between two states, characterised by a junctional cortical and an apicomedial actin network. Our results highlight that behavioural change relies on tightly controlled cellular contractility. Moreover, we show that constriction can occur without pulsing, raising questions why constricting cells pulse in some contexts but not in others.

Morpholinos: studying gene function in the chick.

The use of morpholinos for perturbing gene function in the chick, Gallus gallus, has led to many important discoveries in developmental biology. This technology makes use of in vivo electroporation, which allows gain and loss of function in a temporally, and spatially controlled manner. Using this method, morpholinos can be transfected into embryonic tissues from early to late developmental stages. In this article, we describe the methods currently used in our laboratory to knock down gene function using morpholinos in vivo. We also detail how morpholinos are used to provide consistency of the results, and describe two protocols to visualise the morpholino after electroporation. In addition, we provide guidance on avoiding potential pitfalls, and suggestions for troubleshooting solutions. These revised techniques provide a practical starting point for investigating gene function in the chick.

Dopamine from the brain promotes spinal motor neuron generation during development and adult regeneration.

Coordinated development of brain stem and spinal target neurons is pivotal for the emergence of a precisely functioning locomotor system. Signals that match the development of these far-apart regions of the central nervous system may be redeployed during spinal cord regeneration. Here we show that descending dopaminergic projections from the brain promote motor neuron generation at the expense of V2 interneurons in the developing zebrafish spinal cord by activating the D4a receptor, which acts on the hedgehog pathway. Inhibiting this essential signal during early neurogenesis leads to a long-lasting reduction of motor neuron numbers and impaired motor responses of free-swimming larvae. Importantly, during successful spinal cord regeneration in adult zebrafish, endogenous dopamine promotes generation of spinal motor neurons, and dopamine agonists augment this process. Hence, we describe a supraspinal control mechanism for the development and regeneration of specific spinal cell types that uses dopamine as a signal.