Histological approaches for high-quality imaging of zooplanktonic organisms Rossana C.N. Melo a, * , Priscila G. Rosa b , Nata ´lia P. Noyma a,b , Wa ˆnia F. Pereira a,b , Luiz E.R. Tavares a , Gleydes G. Parreira c , He ´lio Chiarini-Garcia c , Fa ´bio Roland b a Laboratory of Cellular Biology, Department of Biology, Federal University of Juiz de Fora, UFJF, Juiz de Fora, MG, Brazil b Laboratory of Aquatic Ecology, Department of Biology, Federal University of Juiz de Fora, UFJF, Juiz de Fora, MG, Brazil c Laboratory of Structural Biology and Reproduction, Department of Morphology, Federal University of Minas Gerais, UFMG, Belo Horizonte, MG, Brazil Received 5 January 2007; received in revised form 3 May 2007; accepted 4 May 2007 Abstract The investigation of the internal organization of zooplankton communities provides important information on the plankton biology with special interest for the study of ecological processes. Zooplanktoners can play a structural function as indicators for ecosystem health or stress, but their study using histological techniques is still limited. Here we report that the internal structure of zooplanktonic organisms can be facilely observed by a histological approach that combines optimal fixation and processing with a plastic resin (glycol methacrylate) embedding, resulting in increased tissue resolution. Using copepods, organisms that can dominate zooplankton assemblages, as models, collected from a tropical ecosystem (Paraibuna river, Brazil), we showed fine histological details of their muscular, nervous and digestive systems, structure of appendages and cell features. Critical advantages of this approach are that it permits optimal preservation and adequate handling of the organisms (embedded in agar after fixation) for further histological processing and investigation. This is important because it prevents both mechanically induced artifacts and loss of these diminutive organisms during the different steps of processing. Moreover, embedding in plastic resin showed a superior imaging of copepod internal structures compared to paraffin embedding. The use of glycol methacrylate is advantageous over paraffin/paraplast embedding by avoiding heat damage, tissue retraction and allowing faster embedding procedure and better tissue resolution. The value of histological approaches in enabling high-quality imaging of the internal structure of copepods is particularly important because these organisms can be used as indicators of environmental changes. # 2007 Elsevier Ltd. All rights reserved. Keywords: Zooplankton; Histology; Light microscopy; Copepods; Glycol methacrilate; Fixation; Fine structure 1. Introduction Zooplankton communities are structural indicators for ecosystem health or stress (Xu et al., 2001). Zooplankton size and biomass have been classically evaluated as a measure of food availability for higher grazers (Melao and Rocha, 2004). Moreover, zooplanktonic organisms pressure all the microbial communities playing a fundamental link for the aquatic trophic cascade (Carpenter and Kitchell, 1996; Melao and Rocha, 2004). Perturbations to zooplankton populations are sufficient to cause quantifiable differences in system-level metabolism. As part of the process to investigate environmental safety of insecticides (Fliedner and Klein, 1996; George et al., 2003; Kreutzweiser et al., 2002; Medina et al., 2004; Van den Brink et al., 2002) or impact of heavy metals (Millward et al., 2001), for instance, microcosm experiments have been conducted to assess the effects of these chemicals on natural zooplankton populations. Selective reduction of zooplankton such as copepods, induced by pesticides, decreases total plankton community respiration, which implicates in significant, concentration-dependent increases in dissolved oxygen and decreases in conductivity among treated enclosures (Kreutzweiser et al., 2004). Although the distribution, taxonomic composition, and size of zooplanktonic assemblages have been extensively studied in www.elsevier.com/locate/micron Micron 38 (2007) 714–721 * Corresponding author at: Universidade Federal de Juiz de Fora, Department of Biology, Campus 36036-900, Juiz de Fora, MG, Brazil. Tel.: +55 32 3229 3206; fax: +55 32 3229 3226. E-mail address: rossana.melo@ufjf.edu.br (R.C.N. Melo). 0968-4328/$ – see front matter # 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.micron.2007.05.002