Those who claim that only the neocortex of the brain is necessary for consciousness sometimes argue that only electrical activity should be taken into account in the definition of death. Finally, it is possible that the criterion for death is the permanent and irreversible loss of cognitive function, as evidenced by the death of the cerebral cortex. All hope of regaining human thought and personality is then gone given current and predictable medical technology. Currently, in most places, the more conservative definition of death – the irreversible cessation of electrical activity throughout the brain, as opposed to the neocortex – has been adopted (e.g., the uniform death determination law in the United States). In 2005, the Terri Schiavo case brought the issue of brain death and artificial feeding to the forefront of American politics. About 500 active collectors of Formica cinerea were hand-selected from each of the three different settlements in the same location (Błędowska Desert). A total of 300 ant larvae (Myrmeleon boron) were collected from the same site. In the laboratory, lion ants were kept in plastic cups half filled with dry sand (7 cm in diameter, 15 cm high), while ants from each colony were kept in separate plastic boxes (25 x 17 x 10 cm) at 24°C and 40-60% RH and 12:12 L:D took place. Cycle. ants were fed a sucrose solution ad libitum, while lion ants were not given any food; Both were allowed to get used to the setup for two days after being transported from the field. Then, on the third day, in each ant colony, four groups of workers were formed (50 ants each) and kept in separate plastic boxes (25 x 17 x 10 cm), while the rest (about 300 per colony) remained in their original plastic boxes.

Two of these groups were untreated controls, and the other two were experimental groups whose life expectancy was artificially shortened by exposing them to ~100% carbon dioxide for 1.5 h. The method used for experimental manipulation has already been used and its effectiveness in shortening the life of ants is well established [15,16]. A control group and an experimental group were used as sources of the ants captured during the tests, and the other two groups of workers were used to determine that the carbon dioxide treatment was successful (checked daily until all were dead). For tests conducted the day after the carbon dioxide treatment, a biological assay for capturing ant larvae was performed in the cups where the ant lions were kept. In each test, a control collector or collector with a reduced life expectancy was deposited in the ants` den. Immediately after the ant was captured, a potential rescuer (one of the remaining control ants of the respective colony) was introduced into the cup, but not into the pit. Each test lasted three minutes, during which it was determined whether rescue behaviour occurred and, if so, the latency up to the first episode of the rescue, the total duration of the rescue and the categories of behaviour indicated by the rescuer. Four categories of rescue behavior were used: shooting at the victim`s limbs/antennae/mandibles, digging around the victim, removing the sand covering the victim, and directly attacking the ant-lion mandibles.

The operational definitions of behavioural categories were the same as in previous studies of ant rescue behaviour [5,6], except that the definition of shooting in this study was applied to antennae and mandibles as well as limbs. The sequence of tests within each colony was balanced for the control and experimental groups. All tests that lasted less than three minutes (because the victim was completely buried under the sand or released from the predator`s grip for some reason) were excluded. The final number of tests in each group of each colony was 30. No ant ants or lions have been used more than once. The data were analysed in SPSS Statistics 21 software (IBM, Warsaw, Poland). The bilateral Fisher Exact Test (FET) was used to detect differences between groups in the occurrence of rescue, pulling, digging, sand removal and ant attack. Data on mortality, latency and rescue time were analyzed using generalized linear mixed models (GLMMs) using a loglink function and a Poisson error distribution. Colony was included as a random factor, while group was used as a fixed factor. Criteria used experimentally to define moribundity and validated as predictive markers of imminent death should be provided (Clarke, 1997).

Unfortunately, these types of observations are often only published in the methods sections of research papers, and therefore many researchers and veterinarians may not encounter or be aware of such refinements. In addition, many researchers may not have a practical appreciation of humans, as opposed to the scientific refinement of an experimental design. Providing simple but practical examples and promoting the dissemination of information on improvements are important aspects of training research personnel in the humane handling of laboratory animals. Assessment of dying disease may be slightly easier in animal populations than in humans. In experimental designs, the exact nature, timing and extent of challenges and subsequent interventions are known and generally standardized. In addition, animal populations can be very homogeneous in terms of age, history, environment and genetic background. Therefore, accurate prediction of imminent death may be more practical in experimental animal models than in human populations. This sequential model was first tested in a kinetic study with lacUV5 and a modified λPR promoter (λPRAL) by comparing the times at which complete and failed syntheses are completed under single-turn conditions. Surprisingly, the aborted synthesis continues long after the synthesis is complete, contradicting the idea that failed transcripts are an unsuccessful precursor. The long-lived transcription complex has been called the moribund complex, thus defined as a complex that produces only aborted and non-complete transcriptions (Kubori and Shimamoto, 1996). The following properties are known for this type of complex.

(i) The moribund complex is formed from the same homogeneous enzyme fraction as the productive complex synthesizing a complete transcript on the λPRAL promoter, and dissociation of the enzyme from the promoter DNA cancels out any differences between these complexes (Kubori and Shimamoto, 1996). (ii) Dying complexes are structurally different from productive complexes: they are plotted slightly backwards at the λPRAL promoter or forward at the T7A1 promoter and have a more exposed conserved region 3 of σ70 (Sen et al., 1998; 2000; Susa et al., 2002). (iii) With several promoters, the condemned complex is transformed into a dead-end complex that still retains short transcripts but has no stretching activity (Kubori and Shimamoto, 1996). Such complexes on the λPRAL promoter are irreversibly blocked at the promoter level as a holoenzyme (Sen et al., 2000). (iv) Initiation to the T7A1 promoter, in which no kinetic evidence of the moribund complex has been provided for a long time, accumulates the moribund complex in a low-salt state, where the initiation mechanism generally branches into productive and non-productive pathways (Susa et al., 2002), and the non-productive pathway may lead to a dead end, as shown in Fig. 1. Therefore, the fate of the dying complex is either inactivation as a dead-end complex or reactivation by transformation into a productive complex, and these transformations occur at different rates depending on the promoter, resulting in variations in promoter characteristics (Kubori and Shimamoto, 1997; Sen et al., 2001; Susa et al., 2002).