High-Dose Ionizing Radiation Impairs Healthy Dendrite Growth in C. elegans

Abstract

Purpose: The nervous system is vulnerable to radiation damage, and further optimization is required to increase the efficacy of radiation therapy while reducing harm to neurons. Given recent developments in heavy ion therapy, experimental models would be valuable to improve these therapies. We used the nematode Caenorhabditis elegans (C. elegans) to evaluate the effects of high-dose radiation on neuron development.

Methods and materials: In this study, we used confocal microscopy to assess dendritic growth of the PVD nociceptor after high-dose gamma-irradiation from a Cs-137 source.

Results: Irradiation during an early larval stage (L2) delayed overall development but also independently impaired dendrite outgrowth in the PVD nociceptive neuron. Irradiation at L4 larval stage did not result in significant alterations in dendrite morphology.

Conclusions: The nematode C. elegans can serve as a high-throughput model to study the effects of high-dose radiation on dendrite growth. We propose that C. elegans can be useful for studies of experimental radiation therapy modalities and dose rates for translational research.

Figure 1

PVD neuron in C. elegans irradiated at larval stage 2 and imaged at larval stage 4. (A) Fluorescent image of larval stage 4 PVD neuron expressing GFP (left) and schematic (right) with labeled PVD soma, axon, and dendritic branches in 0 Gy control. (B-D) Fluorescent images showing PVD at larval stage 4 after irradiation in the larval stage 2 stage at (B) 0 Gy (sham), (C) 10 Gy, (D) 20 Gy, and (E) 40 Gy. (F) PVD dendrite branch count by dose in larval stage 4 larvae (mean ± SEM). Individual data points correspond to branch number measured in a single animal. PVD 4⁰ branches show a dose-dependent decrease. 2-way analysis of variance corrected for multiple comparisons using Sidak test. Alpha 0.05 (95% CI) P < .0001 (****). N = 6-9 per dose group.

Figure 2

High dose radiation (40 Gy) delays C. elegans larval development. (A) Representative images of vulval substages 4.7 to 4.9. (B) Individual data points represent vulval-substages (4.0-4.9) during larval stage 4 development. Sham (nonirradiated) controls and larvae exposed to 40 Gy at the larval stage 2 larval stage were scored for vulval morphology after 49 hours of growth at 20°C. Vulval morphogenesis was significantly retarded in 40 Gy-treated versus control samples. Unpaired t test (parametric, 2-tailed). Alpha 0.05 (95% CI), P < .0001 (****). N = 20 per dose group.

Figure 3

Irradiation during larval development impairs PVD dendritic branching. (A-B) Fluorescent images showing representative PVD neurons in larval stage 4 larvae after irradiation at the larval stage 2 stage with either (B) 0 Gy (sham) or (B) 40 Gy. (C) PVD dendrite branch count by dose (mean ± SEM). Individual data points represent the number of each type of branch in a single animal. Vulval morphology was used to age-match sham and 40 Gy treatment groups. PVD quaternary (4^o) branching is reduced at 40 Gy in comparison to control. Multiple unpaired t tests (parametric, 2-tailed). Alpha 0.05 (95% CI), ns (not significant), P < .0001 (****). N = 10 per dose group.

Figure 4

Irradiation of the mature PVD neuron at the larval stage 4 does not result in dendritic branching defects. (A-B) Fluorescent images showing representative PVD neurons in adults after irradiation with (A) 0 Gy (sham) or (B) 20 Gy at the larval stage 4. (C) PVD dendrite branch count by dose (mean ± SEM). Individual data points represent the number of each type of PVD branch (2^o, 3^o, 4^o) in a single animal. Multiple unpaired t tests (parametric, 2-tailed). Alpha 0.05 (95% CI), ns (not significant). N = 9 (sham) and N = 15 (40 Gy).