Sex Influence on the Formation of Alcohol Preference and Behavior in Rats during the Long-Term Caffeine and Ethanol Intake

Last years the intake of energy drinks (drinks containing caffeine and alcohol) both in young men and women has been increased. However the investigators do not pay attention to the analysis of gender differences. The goal of the study was the investigation of ethanol and caffeine influence on the alcohol preference formation and behavior in male and female rats under the conditions of long-term experiment. It has been found that six month intake of caffeine, ethanol and their combination has led to the increase of alcohol preference both in male and female rats. Alcohol preference was formed earlier in rats consumed combination of caffeine with ethanol, later on in rats consumed ethanol. In animals consumed caffeine the strong alcohol preference did not form up to the end of the experiment. Alcohol preference was higher in female rats consuming caffeine with ethanol and pure ethanol solution compared to male rats. .Behavioral activity significantly increased in females consumed caffeine and caffeine with ethanol, compared to animals received ethanol and controls. Similar tendency was observed in male rats. The anxiety level was significantly higher in females rats in all experimental groups compared to controls, while males did not demonstrate increased anxiety.

Genetic mutation of Tas2r104/Tas2r105/Tas2r114 cluster leads to a loss of taste perception to denatonium benzoate and cucurbitacin B

Background:Bitter taste receptors(Tas2rs)are generally considered to sense various bitter compounds to escape the intake of toxic substances.Bitter taste receptors have been found to widely express in extraoral tissues and have important physiological functions outside the gustatory system in vivo.Methods:To investigate the physiological functions of the bitter taste receptor cluster Tas2r106/Tas2r104/Tas2r105/Tas2r114 in lingual and extraoral tissues,multiple Tas2rs mutant mice and Gnat3 were produced using CRISPR/Cas9 gene-editing technique.A mixture containing Cas9 and sgRNA mRNAs for Tas2rs and Gnat3 gene was microinjected into the cytoplasm of the zygotes.Then,T7EN1 assays and sequencing were used to screen genetic mutation at the target sites in founder mice.Quantitative real-time polymerase chain reaction(qRT-PCR)and immunostaining were used to study the expression level of taste signaling cascade and bitter taste receptor in taste buds.Perception to taste substance was also studied using twobottle preference tests.Results:We successfully produced several Tas2rs and Gnat3 mutant mice using the CRISPR/Cas9 technique.Immunostaining results showed that the expression of GNAT3 and PLCB2 was not altered in Tas2rs mutant mice.But qRT-PCR results revealed the changed expression profile of m Tas2rs gene in taste buds of these mutant mice.With two-bottle preference tests,these mutant mice eliminate responses to cycloheximide due to genetic mutation of Tas2r105.In addition,these mutant mice showed a loss of taste perception to quinine dihydrochloride,denatonium benzoate,and cucurbitacin B(CuB).Gnat3-mediated taste receptor and its signal pathway contribute to CuB perception.Conclusions:These findings implied that these mutant mice would be a valuable means to understand the biological functions of TAS2Rs in extraoral tissues and investigate bitter compound-induced responses mediated by these TAS2Rs in many extraoral tissues.

A novel quantified bitterness evaluation model for traditional Chinese herbs based on an animal ethology principle

Traditional Chinese herbs(TCH) are currently gaining attention in disease prevention and health care plans. However, their general bitter taste hinders their use. Despite the development of a variety of taste evaluation methods, it is still a major challenge to establish a quantitative detection technique that is objective, authentic and sensitive. Based on the two-bottle preference test(TBP), we proposed a novel quantitative strategy using a standardized animal test and a unified quantitative benchmark. To reduce the difference of results, the methodology of TBP was optimized. The relationship between the concentration of quinine and animal preference index(PI) was obtained. Then the PI of TCH was measured through TBP, and bitterness results were converted into a unified numerical system using the relationship of concentration and PI. To verify the authenticity and sensitivity of quantified results,human sensory testing and electronic tongue testing were applied. The quantified results showed a good discrimination ability. For example, the bitterness of Coptidis Rhizoma was equal to 0.0579 mg/mL quinine, and Nelumbinis Folium was equal to 0.0001 mg/mL. The validation results proved that the newassessment method for TCH was objective and reliable. In conclusion, this study provides an option for the quantification of bitterness and the evaluation of taste masking effects.