Marty Shankland

Segmental specificity and lateral asymmetry in the differentiation of developmentally homologous neurons during leech embryogenesis

This paper describes the embryonic development of three leech neurons which undergo spatially regulated patterns of differentiation. In leeches, the nervous system arises from an iterated array of embryonic cell lineages, and each neuron is represented by a set of bilaterally symmetric and segmentally repeated homologs. Two of the cells discussed here, the neurons nz4 and mz3, stain with antibodies to the neuropeptides SCP and FMRFamide during the course of their embryonic differentiation, but only a subset of the initially immunoreactive homologs continue to express this immunoreactivity into postembryonic life. Those nz4 cells which retain immunoreactivity are referred to as RAS neurons, and the persistently immunoreactive mz3 cells referred to as CAS neurons. The subset of homologs which show persistent expression is segment specific, such that the mature RAS and CAS neurons occupy different segmental domains. In addition, both neurons display a final pattern of expression which is laterally asymmetric, with only one of the two homologs in each segment maintaining the RAS or CAS phenotype. Asymmetric differentiation can occur in either orientation for any given segment, although there is a very strong tendency for the persistently immunoreactive cells to lie on opposite sides of successive segments. The fate of the transiently immunoreactive homologs is unclear, but labeling with intracellular lineage tracers suggests that there are some mz3 neurons which survive late into postemobryonic life and never express detectable levels of immunoreactivity. Intracellular lineage tracers also allowed us to follow the development of a third neuron, mz4, which does not stain for either peptide. The mz4 neuron is initially paired, but undergoes an asymmetric pattern of cell death which also shows a strong tendency to alternate sides in successive segments. These spatially coordinated patterns of neuronal survival and/or differentiation suggest that cell interactions play a role in determining the developmental choices made by individual neurons, and a subsequent paper will characterize those interactions through experimental manipulation.

Developmental origin of segmental differences in the leech ectoderm: Survival and differentiation of the distal tubule cell is determined by the host segment☆

The body plan of the adult leech is metameric, with each hemisegmental complement of ectodermal and mesodermal tissues being produced from a set of seven serially repeated embryonic blast cells. Previous studies have shown that homologous o blast cells give rise to an almost identical complement of descendant cells in each of the 21 abdominal segments, but that one o blast cell derivative--the distalmost cell of the nephridial tubule--is only present in 15 abdominal segments in the mature leech. Here we show that all o blast cells generate a presumptive distal tubule cell and that this cell migrates to its normal position in all abdominal segments. However, in segments which normally do not contain the mesodermal portion of the nephridium, the distal tubule cell dies before undergoing its terminal morphological differentiation. To ascertain whether the fate of the distal tubule cell is determined by its lineage history or by the segmental environment into which it is born, we utilized a previously described procedure for altering the segmental register between different embryonic cell lines. This procedure allowed us to effectively transplant o blast cells into more posterior segments prior to the cell divisions which generate their descendant clones. The results indicate that the survival or death of the distal tubule cell is determined by the identity of the host segment and that a given distal tubule cell could be effectively murdered or rescued by slipping its blast cell precursor into an appropriate segment. These findings suggest that the segment-specific pattern of distal tubule cell survival is not inherent to the O cell line, but arises from interactions with surrounding tissues.

Neuronal competition determines the spatial pattern of neuropeptide expression by identified neurons of the leech

Staining adult and embryonic leech ventral nerve cords with antibodies raised against the molluscan neuropeptides small cardioactive peptide B (SCP) and FMRFamide results in segment-specific and bilaterally asymmetric patterns of cell staining. One immunoreactive neuron, the RAS interneuron, is present in only four rostral segmental ganglia, while another, the CAS interneuron, is restricted to the four most caudal abdominal ganglia and tail. In addition to their segment-specific distributions, only one RAS or CAS cell is found in each segmental ganglion, and they alternate sides between adjacent ganglia (either L-R-L-R or R-L-R-L) with a fidelity of about 95%. This paper utilizes cell deletion techniques to investigate the determination of the asymmetric and alternating pattern of RAS and CAS neurons. We show that developmentally equivalent RAS and CAS homologs are present on both sides of the appropriate ganglia, and that within each ganglion one of the initially paired homologs loses the ability to assume the immunoreactive RAS or CAS fate 2-3 days after axonogenesis has begun. These experiments suggest that there is a competitive interaction between bilateral homologs which ensures that only one mature RAS/CAS neuron is formed per ganglion, and that contralateral RAS/CAS neurons are not required in the same or adjacent ganglia for the determination of the RAS or CAS developmental pathways. Nerve cord transections between ganglia in the CAS domain can alter the spatial pattern of CAS neuron determination, confirming that both bilateral homologs retain the ability to express neuropeptide until late embryonic stages, and suggesting that the alternating pattern of RAS/CAS cells requires communication between adjacent ganglia through the longitudinal connectives.

Cell lineage in leech embryogenesis

Abstract The leech embryo develops into its mature form through an essentially invariant sequence of cell lineages. Individual cell divisions are associated with specific determinative events, but experimental studies indicate that some divisions have several potential outcomes whose selection is under the influence of cell interactions. Present work focuses on the way in which cell divisions and interactions become organized into a complete developmental pathway.

Chapter 2 Position-Dependent Cell Interactions and Commitments in the Formation of the Leech Nervous System

Publisher Summary This chapter discusses the cellular basis of a particular instance of positional specification—the divergence of the O and P cell lines that occur during the embryonic development of the leech. Positional specification requires that cells be able to detect and respond to positional cues by choosing one of two or more alternative developmental pathways. The means by which cells detect and respond to position is a central issue in the field of pattern formation, and by studying this process within a context of defined cell lineages, it may be possible to resolve positional cues in terms of specific interactions occurring among a manageable number of cells. The chapter describes the way in which position-dependent cell interactions become converted into cell-intrinsic states of commitment, leading to a discussion of the ways in which developmental commitment of an embryonic progenitor cell may influence the composition of its descendant clone. Position-dependent cell interactions are a fundamental part of vertebrate neurogenesis and an understanding of pattern formation in invertebrate nervous systems may lend insights there as well.

Segmental diversification of an identified leech neuron correlates with the segmental domain in which it expresses lox2, a member of the hox gene family

The cellular colocalization of LOX2 protein and small cardioactive peptide (SCP)-like immunoreactivity was studied in the nerve cord of the glossiphoniid leech Helobdella triserialis. Of the six neurons that express SCP in the midbody segments 7 to 17, only one, the MPS neuron, expresses LOX2 protein. The medial paired SCP (MPS) neurons are segmentally repeated and can be divided into three contiguous segmental domains according to cell body size and the timing and level of SCP expression. MPS neurons located in the anterior and middle segmental domains express LOX2 protein. In the middle domain, large MPS neurons begin to accumulate SCP shortly after the end of embryonic development, whereas in the anterior domain the MPS neurons are smaller and begin to express SCP at a later stage. In the posterior domain the MPS neurons exhibit a third phenotype -- they have large cell bodies, express low levels of SCP starting from the midjuvenile stage, and do not show detectable LOX2 expression. Lineage tracer injections showed that the MPS neurons arise from a stereotyped cell lineage and are descended from the O teloblast stem cell. In midbody ganglia 2 to 6 and 18 to 21, there are lineally homologous neurons that do not express either LOX2 protein or SCP. Thus, the boundaries of LOX2 expression coincide precisely with two of the segmental boundaries of MPS differentiation, suggesting that expression of LOX2 at the level of this single identified neuron governs some, but not all, aspects of the neurons segmental diversification.