Morphological Characterization of Thoracic Paravertebral Post-Ganglionic Neurons during Development in the Mouse Open Access

Makhlouta Lugo, Camila Sofia (Spring 2020)

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The sympathetic nervous system represents one major system involved in the autonomic control of body function. The entire central sympathetic output arises from spinal cord pre-ganglionic neurons located in thoracic and upper lumbar spinal segments that project to sympathetic post-ganglionic neurons (SPNs). SPNs innervate target organ systems. Most are embedded in paravertebral sympathetic chain ganglia that provide broad distributed control of cardiovascular and thermoregulatory systems. Very little is known about the morphology and plasticity of SPNs in thoracic chain ganglia across post-natal development. Furthermore, few studies have been conducted in mice. This thesis undertook detailed morphological characterization of individually-reconstructed SPNs in the mouse fifth thoracic ganglia at three key developmental time points; weaning (PND 21), adolescence (PND 35) and adulthood (PND 60+). To visualize individual SPNs for unbiased morphological reconstruction, I used sparse labeling tyrosine hydroxylase (TH)-Cre :: tdTomato-reporter mice that resulted in stochastic recombination-based labeling in ~6% of SPNs. I compared developmental changes in cell soma volume and several dendritic properties. Surprisingly, no age-dependent changes in cell volume was seen, though there was significant inter-age variability (at PND 21 and 60+). PND 21 SPNs had the highest number of primary dendrites. SPNs possessed branching and un-branching dendrites. PND 35 SPNs had the highest and PND 60+ had the lowest fraction of branching dendrites. Primary dendrite length of branching dendrites was shortest at PND 21 while PND 60+ SPNs had the longest un-branching dendrites. Overall results support a prominent age-dependent lengthening of dendrites. An unanticipated observation was the immense variation in morphology seen between mice of the same age. This may reflect differences in subpopulation of SPNs sampled by stochastic sparse labeling. Additionally, as considerable SPN connectivity occurs post-natally, variation in early-life experience may also contribute to inter-animal variability. In summary, unbiased labeling identified SPNs as possessing a broad range of morphological variability but with classifiable differences in dendritic properties during post-natal developmental. By using mice as the de facto genetic mammalian model, this work helps lay the foundation for future studies on SPN organization and plasticity including in disease models of autonomic dysfunction.

Table of Contents

1.     Introduction and Background. 1

1.1        Overview. 1

1.2        The Sympathetic Nervous System. 2

1.3        Functional Implications of Morphological Measures. 5

1.4        Development of Sympathetic Chain Ganglia Neurons. 6

1.5        Hypothesis. 10

2.     Materials and methods. 10

2.1        Subjects. 10

2.2        Age Categories: Developmental Aspect. 14

2.3        Tissue Preparation, Imaging and Tracing. 14

2.4        Data Analysis. 18

3.     Results. 19

3.1        Neuron Number Across Development. 19

3.2        Primary Dendrite Number Across Development. 20

3.3        Comparing Incidence of Primary Dendrite Branching Across Development. 22

3.4        Comparing Dendrite Length Across Development. 24

3.4        Cell Body Volume Across Development. 28

3.5        Rostro-caudal Analysis. 32

3.6        TH+ and Td+ Cell counts. 34

4.     Conclusion. 35

5.     Discussion. 37

5.1        Implications of Morphological Trends: Dendritic Arborization. 37

5.2        Implications of Morphological Trends: Cell size. 40

5.3        Individual and Rostro-caudal Variability. 41

5.5        The Sparse Label 43

5.6        Body Size as a Confounding Variable. 46

6.           References 46

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