Exposure time to caffeine affects heartbeat and cell damage‐related gene expression of zebrafish Danio rerio embryos at early developmental stages

Abstract

Caffeine is white crystalline xanthine alkaloid that is naturally found in some plants and can be produced synthetically. It has various biological effects, especially during pregnancy and lactation. We studied the effect of caffeine on heartbeat, survival and the expression of cell damage related genes, including oxidative stress (HSP70), mitochondrial metabolism (Cyclin G1) and apoptosis (Bax and Bcl2), at early developmental stages of zebrafish embryos. We used 100 µm concentration based on the absence of locomotor effects. Neither significant mortality nor morphological changes were detected. We monitored hatching at 48 h post-fertilization (hpf) to 96 hpf. At 60 and 72 hpf, hatching decreased significantly (P < 0.05); however, the overall hatching rate at 96 hpf was 94% in control and 93% in caffeine treatment with no significant difference (P > 0.05). Heartbeats per minute were 110, 110 and 112 in control at 48, 72 and 96 hpf, respectively. Caffeine significantly increased heartbeat - 122 and 136 at 72 and 96 hpf, respectively. Quantitative RT-PCR showed significant up-regulation after caffeine exposure in HSP70 at 72 hpf; in Cyclin G1 at 24, 48 and 72 hpf; and in Bax at 48 and 72 hpf. Significant down-regulation was found in Bcl2 at 48 and 72 hpf. The Bax/Bcl2 ratio increased significantly at 48 and 72 hpf. We conclude that increasing exposure time to caffeine stimulates oxidative stress and may trigger apoptosis via a mitochondrial-dependent pathway. Also caffeine increases heartbeat from early phases of development without affecting the morphology and survival but delays hatching. Use of caffeine during pregnancy and lactation may harm the fetus by affecting the expression of cell-damage related genes.

Gut microbiota mediate caffeine detoxification in the primary insect pest of coffee

“…Caffeine imparts a bitter taste that inhibits insect feeding 6 7 ; it also paralyses and intoxicates insects by inhibiting phosphodiesterase activity and increasing the intracellular levels of cyclic AMP 5 8 . Caffeine toxicity is also related to its negative effects on DNA repair and recombination pathways, and the delay of cell cycle progression as shown in systems such as yeast and zebrafish 9 10 . Because of its demonstrated negative effects on insects, arachnids, slugs and snails, caffeine is considered a natural pest repellent 5 11 12 and, while over 850 insect species can feed on other parts of the coffee plant 13 , and a few of them occasionally on the coffee seed, only Hypothenemus hampei (Ferrari) (Coleoptera; Curculionidae: Scolytinae) has developed the ability to feed and complete its life cycle solely on the economically important caffeine-rich coffee bean.…”

Zebrafish Model for Safety and Toxicity Testing of Nutraceuticals

“…Capiotti et al (2011) evaluated the effect of caffeine on zebrafish development. The effect of caffeine on heartbeat, survival, and expression of cell damage-related genes was reported by Abdelkader et al (2013). Another study on caffeine was performed during zebrafish embryonic development.…”

“…They suggested that caffeine was able to affect the expression pattern of cellular target genes in the early stages of zebrafish development, but (2008) . Abdelkader et al (2013) concluded that it may harm the fetus if it is used during pregnancy and lactation. A green fluorescent vascular endothelium transgenic line was used to assess the induced toxicity of caffeine during embryonic vascular development, but no defects were found in vasculogenesis; however, real-time PCR showed that caffeine treatment can disturb the angiogenesis of zebrafish embryos (Yeh et al, 2012).…”

Deep hypometabolic state mitigates radiation-induced lethality in zebrafish

“…Radioprotection or mitigation due to DH state was calculated by comparing the DH + radiation (DH + IR) group with the radiationalone group. Quantitative analysis of heart rate was done following the stopwatch method [26,27]. Briefly, embryos were anesthetized in 0.001% tricaine dissolved in 1× E3 medium prior to counting under a microscope (Leica DMR, Germany) connected with a computer by an external camera (MDCE-5C, USB 2.0).…”