Discrete Targeting of Peptide and Non-Peptide Neurotransmitters by a Single Neuron

Date of Award


Degree Name

Doctor of Philosophy


Biological Sciences

First Advisor

Dr. John Jellies

Second Advisor

Dr. Christine Byrd-Jacobs

Third Advisor

Dr. William F. Jackson

Fourth Advisor

Dr. Cindy Linn


Hirudo, acetylcholine, FMRFamide, discrete targeting, heart tube, varicosities


The heart excitor (HE) motor neuron in the medicinal leech, Hirudo, releases acetylcholine (ACh) and a peptide, FMRFamide, to regulate the contractile activity of the heart tube and associated side vessels. Consistent with Dale’s Principle, it was assumed that both neurotransmitters were coreleased by all presynaptic varicosities. However, discrete peptide-positive and peptide-negative varicosities can be found consistently at particular sites of innervation on the heart tube. Dual-labeled HE neurons were produced by pressure injecting Neurobiotin into single HE neurons and applied antibodies against FMRFamide on the same preparations. Consistent with initial expectations, peptide-labeled varicosities were numerous and widely distributed along the heart tube and at one of the three side vessels, the lateroabdominal vessel (LaV). Nevertheless, some Neurobiotin-labeled varicosities along the heart tube lacked peptide label entirely. Moreover, there were dense and distinct peptide-negative clusters of varicosities at the valve junctions of the other two afferent side vessels at each segment, the latero-dorsal vessels (LdV), and laterolateral vessels (LlV). The peptide label was, however, found in HE axons and varicosities that projected distally along the two afferent side vessels. Therefore, the more proximal peptide-negative clusters could not simply be the result of deficient staining of peptide. Rather, it is inferred that FMRFamide is transported (or excluded) to specific locations on the heart tube and that ACh is present in varicosities that lacked peptide. Furthermore, the development of these peptide-negative varicosities can occur earlier than the development of peptide-positive varicosities and is independent of the postsynaptic activity of the heart tube that was entrained by the central pattern generator. Nevertheless, since rhythmic heart tube activity was still present, it strongly suggests that the development of peptide-positive varicosities must occur during a critical period of activity-dependent development. Taken together, the discrete targeting of neurotransmitters to discrete sites on the heart tube as well as the sequential development of corelease of neurotransmitters by a single neuron can be explained using a discrete targeting model of synaptic transmission. Compared to Dale’s Principle, this model is an alternative that provides a more nuanced perspective of synaptic transmission than previously understood.


Fifth Advisor: Dr. John Spitsbergen

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