Constipation (animal) study model design includes following headlines:
Species : Mus musculus (Mouse)
Strain : C57BL/6J mice, Balb/c or Swiss albino
Age : 3-4 weeks
Body Wight : 20-25 g
Sex : Male
No. of animals : 10 /Group (20 only for testing group-Natural Probiotic Formulation)
Total Animals : 70
2.0 ALLOCATION OF GROUPS:
|
Groups |
Treatment |
Dose; ROA* |
No. of Animals |
|
G1 |
Saline control |
15-20 ℃ saline-treated p.o. |
10 |
|
G2 |
Ice-cold
group |
0-4 ℃ saline-treated p.o. |
10 |
|
G3 |
Ice-cold
group (treated) |
0-4 ℃ saline-treated p.o. + NP-1 |
20 |
|
G4 |
Ice-cold
group (treated) |
0-4 ℃ saline-treated p.o.+ NP-2 |
20 |
|
G5 |
Ice-cold group (Standard) |
0-4 ℃ saline-treated p.o.+ Standard |
10 |
2.0 METHOD:
· To eliminate the influence of the biological rhythms, intragastric administrations are conducted at 8:00 am once daily for 14 days.
· Mice in ice-cold (G2-G5) and (G1) saline control groups are initially administered either ice-cold or room-temperature saline, respectively, at a dose of 0.2 mL/mouse, and then the dose should be increased by 0.05 mL/mouse every 5 days.
· Animals are normally raised and monitored for an additional 6 day after the cessation of the intragastric administration period after this, stool is collected with ceramic cups.
· After collection, the changes in stool form (pellet size) are observed and recorded before weighing. Then, each ceramic cup weighs to determine wet weight using an electronic balance.
· After drying in a microwave oven (medium heat) for 6 min, the cup is weighed again to determine the dry weight. After the completion of treatment, the differences between the five groups are evaluated at 15 days and 20 days.
· Mice in each group are randomly assigned to a 15 days subgroup and a 20 days subgroup. The 15 days subgroup is used in the intestinal propulsion experiments at 15 days while the 20 days subgroup is raised normally and used in the same experiments at 20 days.
· After 12 h of fasting, mice are intragastrically administrated a suspension of black carbon (0.3 mL) and killed 10 min later via cervical dislocation. Then, the section extending from the pylorus to the ileocecal valve is removed.
· The full length of the intestinal tract as well as the propulsive distance of black carbon in the tract is measured under a tension-free state, and the ratio of the propulsive distance to the length of the intestinal tract is measured in all groups.
· Additionally, in the next experiment at the 15 days and 20 days time points, we also determined the defecation time after similarly dividing the mice into the 15 days and 20 days subgroups.
· After 12 h of fasting, the red carbon suspension (0.3 mL) is administered, and the time required to defecate the first stool pellet containing the red indicator is recorded.
· Animals administered prokinetic agents are pretreated with domperidone suspensions (0.3 mL) for 4 h. Tissue preparation & Immunohistochemistry with help of primary antibodies for PGP9.5.
3.0 END POINT PARAMETER(S):
· Stool collection and examination
· Measurement of intestinal function
· Feed water consumption
· Body Weight.
· Tissue preparation
· Immunohistochemistry
4.0 REFERENCE(S):
· Chao Liang, Kai-Yue Wang, Zhi Yu, Bin Xu. Development of a novel mouse constipation model. World J Gastroenterol 2016 March 7; 22(9): 2799-2810. DOI: 10.3748/wjg.v22.i9.2799
· Ji Eun Kim, Jun Go, Ji Eun Sung, Hyun Ah Lee, Woo Bin Yun, Jin Tae Hong and Dae Youn Hwang. Uridine stimulate laxative effect in the loperamide-induced constipation of SD rats through regulation of the mAChRs signaling pathway and mucin secretion BMC Gastroenterology (2017) 17:21 DOI 10.1186/s12876-017-0576-y
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