My thesis research focused on the neural mechanisms underlying the crawling locomotor behavior in the medicinal leech Hirudo verbana at the levels of the neural network and individual neuron. Crawling in the leech was studied because it possesses great flexibility in its movements but is simpler than walking, especially in vertebrates. In Chapter 2, I provided the first evidence that dopamine (DA) promotes crawling in intact but isolated CNS preparations. I verified the rhythmic activity induced by DA was authentic crawling by monitoring key crawl-related motoneurons that are active during overt crawling. I established that DA-induced bursting propagated along the CNS in a metachronal wave moving from anterior to posterior. I also showed that DA activated the full complement of both excitatory and inhibitory motoneurons in single isolated ganglia located in all regions of the CNS. These results established that each segmental division of the CNS contains a complete set of rhythm-generating elements (i.e., a 'unit burst generator') for crawling. My results, coupled with DA's ability to terminate fictive swimming (Crisp and Mesce, 2004), also elucidated the role of DA potentially acting as a chemical decision-making cue.
During complex locomotor behaviors like walking or crawling, precise timing and coordination of motoneuron activation and deactivation must occur both within and between segments, while still maintaining sufficient flexibility of the behavioral outputs. Chapter 3 investigated the mechanisms in the CNS underlying the coordination of the segmental neural oscillators for crawling. I determined that descending signals from the cephalic ganglion (i.e., brain) are necessary and sufficient for the expression and intersegmental coordination of crawling. I also determined that each segmental crawl oscillator communicates an excitatory 'crawl-like' drive to nearby oscillators, via the hemi-connectives, with the strongest communication occurring toward the posterior adjacent ganglion. Importantly, this crawl-like drive is not sufficient for generating normal intersegmental coordination. My results provide evidence that both long-distance descending and local inter-oscillator coupling contribute to crawling. This dual contribution helps to explain the inherent flexibility of crawling, and provides a foundation for understanding other dynamic locomotor behaviors across animal groups.
In Chapter 4 I posed the over-arching question: What contributions can a single neuron have on controlling the behavior of an animal? To address this important question, I studied R3b-1, a long-distance projecting neuron that is bilaterally paired and located in the cephalic ganglion of the medicinal leech. I established that this interneuron has the cytoarchitecture to communicate potentially with both the left and right halves of each segmental ganglion within the CNS. I also determined that R3b-1 activity is particularly important for the expression of fictive crawling, as spiking in a single R3b-1 was determined to be both necessary and sufficient for crawling behavior. Aside from its command-like properties, R3b-1 was found to modulate the frequency of crawl-related motor outputs; furthermore, when the CNS was biased to crawl by exposure to DA, crawling became the exclusive locomotor pattern produced by R3b-1 as opposed to a mixture of crawling and swimming outputs. Although the above attributes render R3b-1 an exceptional cell, it is its ability to coordinate the segmentally-distributed crawl oscillators that makes this singular neuron so notable. To my knowledge, this cell provides the first biological example of a single command-like neuron that is also vital for the intersegmental coordination of a locomotor behavior.
During my dissertation research I made a number of discoveries that have advanced our knowledge of the underlying organization and cellular mechanisms that generate crawling in the medicinal leech. My discoveries add to an already vast corpus of information about the neural underpinnings of motor behaviors in the leech and also shed light on potential mechanisms that may generate flexible locomotion in other animals such as walking vertebrates. Ultimately, my research may advance our overall understanding of complex control schemes that govern rhythmic behaviors in general.