SpotMetrics: An Open-Source Image-Analysis Software Plugin for Automatic Chromatophore Detection and Measurement.

Coleoid cephalopods (squid, octopus, and sepia) are renowned for their elaborate body patterning capabilities, which are employed for camouflage or communication. The specific chromatic appearance of a cephalopod, at any given moment, is a direct result of the combined action of their intradermal pigmented chromatophore organs and reflecting cells. Therefore, a lot can be learned about the cephalopod coloration system by video recording and analyzing the activation of individual chromatophores in time. The fact that adult cephalopods have small chromatophores, up to several hundred thousand in number, makes measurement and analysis over several seconds a difficult task. However, current advancements in videography enable high-resolution and high framerate recording, which can be used to record chromatophore activity in more detail and accuracy in both space and time domains. In turn, the additional pixel information and extra frames per video from such recordings result in large video files of several gigabytes, even when the recording spans only few minutes. We created a software plugin, "SpotMetrics," that can automatically analyze high resolution, high framerate video of chromatophore organ activation in time. This image analysis software can track hundreds of individual chromatophores over several hundred frames to provide measurements of size and color. This software may also be used to measure differences in chromatophore activation during different behaviors which will contribute to our understanding of the cephalopod sensorimotor integration system. In addition, this software can potentially be utilized to detect numbers of round objects and size changes in time, such as eye pupil size or number of bacteria in a sample. Thus, we are making this software plugin freely available as open-source because we believe it will be of benefit to other colleagues both in the cephalopod biology field and also within other disciplines.