Mechanisms of rhythm generation in the lamprey locomotor network

Animals propel themselves through space, swimming, walking or flying, by rhythmic oscillatory movements of their body and limbs. In vertebrates these movements are mainly generated by specialized neuronal circuits in the spinal cord, called central pattern generators (CPGs) for locomotion.

Spinal networks integrate sensory feedback about body position and the environment, together with supra-spinal motor commands, to produce precise and goal-directed locomotion. The lamprey, an aquatic vertebrate that swims by rhythmic left-right bending of its body, is an advantageous system for the study of CPGs. It conserves ancient features of the vertebrate nervous system, with a lower complexity and limited number neurons. Its isolated spinal cord, when activated pharmacologically, produces at the ventral roots the rhythmic motor pattern of swimming.

This allows the experimenter to investigate in vitro, the neuronal mechanisms responsible for locomotion. Individual neurons or pairs of a pre- and a postsynaptic neuron can be recorded from, to study their cellular and synaptic properties and gain an understanding of how different cells are wired together in a network. The work presented in this thesis aims at clarifying three crucial aspects of how the locomotor rhythmicity arises.

The core of the CPG is considered to be organized in two populations of excitatory interneurons (EINs), one on the left and one on the right side of the spinal cord, each providing phasic glutamatergic excitation to the ipsilateral motoneurons (MNs). Moreover, these two groups of EINs inhibit one another by way of interposed glycinergic interneurons with crossed projections, ensuring a proper alternation of activity. It has been proposed that this crossed inhibition plays an essential role, not only in the left-right alternation, but also in the generation of rhythmic bursting.

We tested this hypothesis by performing complete midline sections of spinal cord pieces and found that the hemicord of the lamprey expresses rhythmic locomotor bursting during pharmacological activation, as well as following electrical stimulation. Thus, the generation of the basic swimming rhythm is provided by unilateral spinal networks in each hemicord, and crossed inhibition is not required for rhythmogenesis. We then explored the firing pattern of single MNs and interneurons during locomotor bursting in the hemicord, and found that active neurons fire one action potential for every locomotor cycle. This has lead to a tentative model for the operation of the unilateral networks in the absence of crossed inhibition.

Among the mechanisms that regulate the frequency of locomotor bursting in the lamprey spinal network, are Ca2+-activated K+ channels (KCa). These open as a consequence of Ca2+ influx during each action potential and generate a slow afterhyperpolarization (sAHP). Summation of the sAHP helps terminate bursts on either side the spinal cord. A crucial question was whether a KCa mediated conductance increase might also take place as a consequence of Ca2+ entry during subthreshold excitatory synaptic input. This would modify the integrative properties of the neuron. We examined this possibility in a classic model of synaptic contact, the giant reticulospinal axon to MN synapse, as well as during the cyclic excitation-inhibition of fictive swimming in MNs. No evidence for a direct interaction between synaptic input and KCa channels was found, thus simplifying dendritic computation.

We also characterized the pharmacology of the sAHP, describing a previously unknown component, mediated by Na+-dependent K+ channels (KNa). We found that during repetitive firing, this component represents a progressively larger proportion of the sAHP. This will make it an important factor contributing to the regulation of the spike frequency of spinal neurons, and potentially also of the locomotor CPG.

List of scientific papers

I. Cangiano L, Grillner S (2003). Fast and slow locomotor burst generation in the hemispinal cord of the lamprey. J Neurophysiol. 89(6): 2931-42. Epub 2003 Jan 22
https://pubmed.ncbi.nlm.nih.gov/12611971

II. Cangiano L, Grillner S (2004). Mechanisms of pattern generation in a spinal locomotor network deprived of crossed inhibition: the lamprey hemicord. [Submitted]

III. Cangiano L, Wallen P, Grillner S (2002). Role of apamin-sensitive k(ca) channels for reticulospinal synaptic transmission to motoneuron and for the afterhyperpolarization. J Neurophysiol. 88(1): 289-99.
https://pubmed.ncbi.nlm.nih.gov/12091554

IV. Wallen P, Cangiano L, Grillner S (2004). Sodium-dependent potassium channels contribute to neuronal frequency regulation. [Submitted]