Rat Handling - Science topic

Hello.

Take alagebrium for example, you can search it on google scholar (or others) and get some articles about it. I picked up one as below.

Am J Transl Res. 2019; 11(3): 1569–1580.; PMCID: PMC6456531

In this article, they mentioned that "......AGEs inhibitor ALT-711 (20 μg/mL; MedChemExpress, China) was added......". So you can turn to "MedChemExpress" to see if the price and distance are ideal or not. If not, turn to other papers.

It has to be mentioned that, you can go to https://scifinder.cas.org/scifinder. Using substance identifier will easily lead you to what you want, though I had not bought through this way.

To me, when thinking about those words and their common uses, I have them divided up in my head a certain way. Your mileage may vary, but here's how I use them.

Acclimation and acclimatization are the same thing-- getting used to life at a different place than the animal producer. I like acclimation better as acclimatization just seems....extra.

Habituation is getting used to something happening--animals are habituated to restraint, they are habituated to a behavioral testing apparatus.

And finally, adaptation is a longer-term event occurring at the species level to permanent changes--the rats are adapted to living at altitude; the mice have adapted to desert life.

If you want to determine feed intake, then pellets are to be preferred. Also then the animals can not select individual feed ingredients.

You can calculate fat content from ingredient composition or determine it chemically.

I have tried facial vein blood collection last Sunday, with 5mm blood collection needle. For mice of 4-5weeks years-old,6-7 drops equal 150-300ul ,then press the wound for 20s. Till now，every thing is ok~

Hi Jesse,

I am from the manufacturer of iPRECIO pumps. I am very late but I thought I would still share. Our pumps will infuse up to 6 months at 1ul/hour. See www.iprecio.com . You can even program the pump to infuse intermittently or to try to simulate the hormonal cycle.

Some references for the choices you mentioned.

Good luck and feel free to contact if you have more questions.

Hello!

I found this article that relates rat age to human's:

Hope it answers your question properly.

Best wishes for your research!

Dear Pradeepkumar.

If you or someone you work with is familiar with pharmacokinetic modelling, you may wish to look into a practice called optimal design. This approach uses certain statistical properties to derive (amongst other possible variables to optimise for) the sampling times at which the maximum amount of information may be retrieved. Especially in studies where the information density may not be easily determined rationally such as interaction studies, this approach may significantly improve the chances of study success. Although it requires investing some time and effort, and some a-priori expected or established model, in my experience it is worth it.

For further reading I suggest the article "Optimal Design of Pharmacokinetic Studies" by Leon Aarons and Kayode Ogungbenro, 2010.

Good luck on your study

Good Question

I believe the acute toxicity of corticosterone in mice is not very high i.e. if you dose only once. So probably the choice of dose is more dependent on what you would like to achieve rather than any limitation by toxicity. Dose ranges in rats should be too far off the ones in mice, maybe those for mice being even a little higher.

The protocols for carrying out similar studies and comparing the data obtained in different laboratories have been developed. Dosages for different substances are chosen, respectively, single, double ...

Try methylene blue. Do not exceed 100-120 µl for adılt mice, and 10 µl for neonates  

Kopf instrument is best device for stereotaxic surgery in Mice

Thank you all! 

Our research team has dealt with this protocol for years. Pilot experiments are needed in order to find the proper combination dosage of STZ and NA, since different results are obtained from lab to lab. Don't be so sure that the achieved diabetes will be the type 2. Also,mind that STZ is photo(light)-sensitive and must be administered freshly after its preparation. Very long fasting  of the animals doesn't help. Not all the animals will be rendered diabetic. In my opinion models combining high-fat diet and (a) low dose(s) of STZ are more appropriate for the study of type 2 diabetes. To my mind, the model you are asking about will easily turn the animals into type 1 diabetes.

look over Modeling bladder cancer in mice: opportunities and challenges - NCBI - National Institutes of Health

Normal rodent islets have tremendous capacity to compensate for hyperglycemia when infused with glucose. Achieving a high blood glucose by infusing glucose solution, comparable to severe hyperglycemia caused by streptozotocin treatment for near complete destruction of beta cells, is almost impossible. 

Ages ago, in case of normal rats (~150gm), I infused 50% glucose at a flow rate of 2ml/hr - within 2 days rats were able to compensate sufficiently that raising blood glucose over 200mg/dl wasn't easy and getting to >400mg/dl was impossible.

Both STZ and Alloxan can be purchased from Sigma Aldrich.

One of the best ways to ensure that the rats consume the high fat powder is to incorporate it into the pellet feed. Adjustments can be made in the relative composition of the pellet to make the high fat powder an integral part.I know from experience that coconut-oil based food may not appeal to rats. In the alternative, consider using vegetable oil; either way, remember to make corrections for the added calories. All the best.

Please check the following link and attachment.

Thanks John

I learned from Harry Broadhurst that cross-fostering was vital for the studies of classical genetics such as development of the Maudsley reactive and non-reactive rats. The main disadvantage was the amount of time and grant money needed to do the extra work. Although I did not engage in genetic studies myself the cross-fostering control was also important for studies of manipulations of maternal history by prenatal exposure to drugs. It was at that point that I realised that countless numbers of studies refer to strain differences in rats without cross-fostering controls and cannot therefore be interpreted as establishing genetic effects. Most people don't seem aware of that. It seems to cast doubt on many studies of genetically manipulated animals (e.g. knock-outs and knock-ins) where cross-fostering appears to be very rarely done. Do the others who have responded to you question agree? If I am right then it is a matter that deserves a public airing.

Hi- If you are going to monitor sanitation of animal house, you can do that according to below methods:

monitoring by using Luminometers (it highly recommended)

Swab Method

RODAC Plate Method

Agar Sausage Method

The Petrifilm Plate Method

all the best

mehran

Dear Anmol, 

In order to calculate the caloric (energy) intake you need to know the energy provided by the rat chow (feed) which your using, e.g. 3020 kcal/kg, (it can be easily available from your laboratory animal chow (feed) supplier. 

After getting this value you have to measure the daily food intake of the individual SD rats.

Caloric Intake(Kcal/g/day) = Daily food intake (gm) * total energy food of (kcal/kg)/1000

From caloric intake you can calculate the Feed Efficiency Ratio

Feed Efficiency Ratio (FER) = Mean Body Weight Gain / Caloric intake

For more insights refer my publication by clicking this link http://authors.elsevier.com/a/1SoS26FNx9S22D

Hope you understand.

Best luck.

You can find the answer in the following research :

Y. Li, J.-G. Wang, Y.-P. Li, Z.-F. Lin, A MODIFIED RAT MODEL FOR CANNULATION AND COLLECTION OF THORACIC DUCT LYMPH. Lymphology 44 (2011) 82-88

You can also contact :

 Zhao-fen Lin, PhD, Professor, Emergency Department

Changzheng Hospital Second Military Medical University

No. 415 Fengyang Road- Shanghai 200003, China

Tel: +8613601605100

E-mail: linzhaofen@yeah.net

"As far as I know, urethane should preferably be adopted in non-recovery procedures,"

This is 100% true, but urethane as "non-recovery" anesthetic usually works relatively safely. However, I made more than 1000 craniotomy on rats, and independently  on the anesthetics, unexpected  deaths always occured. In my lab, the case was usually something pulmonary problem.  My preferable drug was always Hypnorm, as operational anesthetic. Subcutaneous injection was the best route of the administration.  

Elevated ACTH but low CORT is sometimes seen in rodent models of post-traumatic stress disorder (only in males though, are you using males? There are significant sex differences in the HPA axis response to stress). Diminished CORT release in response to stress could be due to a sensitization (or increase in number) of glucocorticoid receptors (GR) in the pituitary, and if all GR are saturated with ligand, no more suppression of ACTH can occur so that's where you can get high ACTH and low CORT. Due to this, the adrenal glands can become insensitive to ACTH--you can quantify ACTH receptors or just give them an ACTH injection and measure their CORT response to see.

I use the single prolonged stress paradigm (a PTSD model), which is a single exposure to 2.5 hours of intense stress. These stressed rats show a 50% increase in glucorcorticoid receptor expression in the PVN of the hypothalamus and exaggerated negative feedback of the HPA axis which drives their CORT levels down. As you pointed out though, CUMS doesn't usually result in this phenotype. It sounds like maybe something was different between your experiments, but it's hard to tell what (light cycle, handling, housing?)

If you want to explore further what's going on in your rats I suggest looking at the dexamathasone suppression test to measure HPA negative feedback (see attached paper). It would also be helpful to quantify glucocorticoid receptor expression in the hypothalamus and pituitary. Baseline CORT levels can vary a lot between individuals, so I take a baseline blood sample and expose them to a mild stress (30 minutes  in a restraint tube), take another blood sample at 30 minutes, and then look at the CORT released. Good luck!

Dear dr , the best method to blood collection in rat is tail vein puncture with insuline syringe but should be veterinarian have experiance do that

I think Apomorphine can induce such licking behaviour.

You might as well ask whether there are differences between between Wistar rats as offered by different breeders or between colonies housed at different universities or research institutes in the world. Yes there are huge differences. And if you order the "same" Wistar rats over the years at the same commercial breeding company, you might get different animals, at least from a behavioural perspective (van der Staay, 1997). Van der Staay also showed huge differences between various outbred rat lines in the size of the consequences of an arterial occlusion.   

Depending on how well the implant is fixed on the skull (screws, how dry the skull was during surgery) you may have the implant stable for over a month. But of course, you are right, astrogliosis and the resulting glial scar is an issue. However, how this will affect neurotransmitter level is probably more dependent on how often you insert the probe into the cannula as this will extend beyond the tip and will disrupt additional ´fresh´ (or less fresh after multiple insertions) tissue. In practice I therefore kept the probe overnight in the cannula before measurements and kept at least a week of recovery in between measurements. Finally, I would not recommend over 4 measurements. You may check effects of repeated insertions on neurotransmitter levels by comparing the baseline throughout multiple measurement-days. 

I think it is difficult to find a "standard" for enrichment of rats. In general at best the distinction between exercise (running wheel) and "enrichment is made. But often these also applied together. In a study(http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0011943) we showed that both forms of enrichment( with and without running wheel) have anxiolytic properties however, there are some studies that show differences between exercise and enrichment (for example : http://learnmem.cshlp.org/content/18/9/605.short

also see: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0016643)

Regards,

Erik

Acute alcohol poisoning, would be my diagnosis....Yellowish fluid in the stomach would be indicative of bile reflux from the duodenum... White fluid would be chyme that got obstructed.. much like a human who is too drunk, bile may build up in the stomach as a acute toxicity response and an anti-peristaltic response of the duodenum in attempt to rid the body of the toxin (i.e. alcohol); but, rats cannot vomit, so the the fluid builds up and the rat will suffer a toxic acute response, based on too much alcohol and stomach contents.. As the duodenum was in reflux mode, it was effectively obstructed so the stomach contents stayed there, not being able to be expelled though vomiting. Alcohol is also well absorbed through the stomach mucosa, so it may be the case your rat died simply of an alcohol overdose associated with acute toxicity from bile and ingested food toxins. As it happened only in one rat may not be surprising; Each individual animal is different, and it is difficult to either monitor exactly the alcohol intake, the food/other intake, the background health status or the susceptibility to alcohol poisoning in each individual.

I agree with the previous answer, except if the volume for the needed active amount of drug exceeds 2ML/kg, which is the maximum volume a rat can receive in a single injection/administration (either ip, sc, iv, or po). This is, at least, according to Swiss regulations which are based on European ones. 

If the volume is not a problem then just inject the fix amount corresponding to the 0.5mM you aim at. The real dose/Kg will depend on the weight of each animal, but maybe that's no a variable of your study...

Good luck,

Daniela

Dear Dr Thoria

I always measure such parameters in my rats "control. However, there is a paper that mentioned that normal serum adinopectin level in control group of rats is 109.15 ± 95.40

Attached please find this paper and you can contact the authors for further support or information.

Sincerely yours

Hi Elcin,

I agree with the precautions proposed by Ignacio.  For the recipe of K : X  cocktail, what I do usually to make as stock solution is simple: I make 50mg/ml ketamine make a 10ml solution, take only 8.8ml of it aside. then make 20mg/ml Xylazine solution (10 ml stock solution) then add 1.2 of the Xylazine solution to the 8.8 ml Ketamine you prepared earlier, the result is 10 ml of cocktail: the working formula for rats ( i.p. injection, and/or i.m. , male or female does not matter):take according to body weight. 2 ul of the cocktail for each gram body weight:

example: rat weighing 210 gms will be given:

210 x 2 = 420ul (0.42 ml) of the cocktail to be injected i.p, or i.m.

230 gm rat (male or female) 230 x 2 = 460 ul (0.46 ml) and so on

350 gm x 2 = 700 ul ie 0.7 ml

the animal should be under anesthesia after 3- 5 minutes and absence of reflexes should last 45 -50 minutes.

one last thing: for storing the cocktail if you have a preservative like benzethonium chloride 0.01% ( or any other ) that's fine, but if you don't, pls make sure you store it at 2-8°C, for not more than a week, otherwise the cocktail will produce inconsistent results.

hope this is helpful, best wishes and good luck.

What is a solvent for Ketamine and Xylazine? - ResearchGate. Available from: https://www.researchgate.net/post/What_is_a_solvent_for_Ketamine_and_Xylazine [accessed Jun 30, 2015].

Apologies, allow me please to clarify the dilemma I am in, when we did OGTT on rats, ( after tedious habitation and handling, we introduced the dosage of glucose load through feeding tube orally, which by itself entails a certain degree of stress to the animal, as some of the blood samples collected are so soon: 15 and 30 min. given the fact that all animals where subjected to the same conditions, one would contemplate that this variable will have an insignificant effect on the results. But, rats are nocturnal creatures, i/e. their most biologically active behavior takes place during the dark cycle ( eating, drinking, playing etc...)  we tried to maximize the minimum stressing conditions in the testing, exploiting the fact they are fasted overnight, by just presenting the glucose load in their drinking bottle. the result where the animals they drink the glucose but not to the correct volume which allows us to be consistent with our  results.

The blood collection is achieved by jugular vein access ( 3 days recovery period after surgery) and the help of the Culex®NxT ABC automated blood sampler. so it is done completely without any interference by the examiner.

The question remains, how to do the oral dosing of the glucose load without stressing the animals?

Thank you again for your interest in helping with this question.

Ghanim.

Hi Nagwa,

yes, you are absolutely correct, it is a very delicate tissue, and the procedure is very delicate as well.

I usually do it under surgical microscope (3D Stereo) with ultrafine microinstruments ( CASTROVIEJO Corneal scissor Angled to side blunt tips, and KRAMER Fixation forceps to get a good atraumatic hold of the eyeball, and CASTROVIEJO straight forceps without teeth to peel off the vitreous body away ) all under the seal of sterile normal saline, if you do not have the proper microsurgical instruments, and proper skills and training, the dissection will not be a successful as you desire.

I looked up an animation for you, I believe it is very descriptive.

best wishes

Hi Marnie

I think you dose is too much decrease the amount, while if you need this such dose as experimental model, I prefer to prepare a mixture of it with Xylazin because it make rats more relax when you are doing experiment

Thanks

Alex, it would depend on their immune status and your experiments. Are they likely to come to harm if housed in normal cages? Would their housing in normal cages cause harm to other animals in the facility?

As Ryan mentioned there will be differences in the animal behaviour based on housing. I wouldn't change the housing in the middle of an experiment. Depending on the nature of your experiment, you may have to repeat earlier experiments if you change the housing.

This article contains lots of information about techinques for blood sampling, including recommended collection volumes and how these vary with frequency of collection. Our vets recommended this paper when we needed to collect repeated, fairly large samples over an extended period. Hope that it helps. 

If you are stressing the rat for a long period of time and food and water is still given freely,you will need to tilt the cage on its  small side.This will allow the rat to drink and eat normally.Our lab we found that the tilting of the cage when we are using mice,is not much of a stressor .But since rats are more sensitive,it may work.

A 'timed pregnant' rat is one in which the day of conception is known, generally because a male breeder is put in with the female at the time at which she is sexually receptive and a sperm plug is then detected. This can be important particularly in cases when embryos are being examined, so that their exact age can be determined. In addition, since pregnancy in rats is lasts between 21 and 23 days, it can allow researchers using birthed offspring to plan their experiments accurately.

Tail vein injection is another alternative. Search "tail vein injection" and you will find considerable information. I became interested in tail vein injections with respect to taste aversion learning. Humans experience vascular taste in that injection of a dilute saccharin solution into the vascular system produces a taste of saccharin in a few seconds when the blood containing saccharin reaches the tongue. Rats apparently experience a comparable taste sensation because tail vein saccharin injection followed by a nausea producing procedure results in a conditioned taste aversion without the rats ever having orally ingested a saccharin taste. 

You can try to use the Clarity technique. If I understood your aim, I think it can work.

These are two useful links you can look at.

Regards

Below is the composition of our pellets for rats. However we do not prepare this ourselves, but buy it from a commercial producer.

Crude Protein : 23 % Crude Fat : 3.0% Crude Fiber : 7.0% Acid Insoluble Ash 8% Calcium : 1-2.5 % Phosphorus : 0.9% Sodium : 0.5-1% Moisture: 12%

Ingredients 
Corn, Soybean pulp, Sunflower seed meal, Shorts, Bonquality flour, Alfalfa pellets, Molases, Meat and bone meal, Poultry meal, Sepiolite, inorganic DCP, Marble dust, vitamins, minerals

In case you use Charles River as your supplier, they have published those data also, even separated for their different breeding facilities. In case simply contact your local CR representative. If you do not use Harlan or Charles River animals, I would suggest to collect as many data as possible from the different breeders and have a look on the variability of the parameters.

calipers, a balance with a container to put the pup into (tare it before adding the pup!) 

practice first - plan well, (ie what you will do with the tissues after will dictate the best way to process/store them)

Hi Thomas - I routinely plate P0 rat hippocampal neurons at both low (40k) and high (>100K) densities to at least DIV21.  I feel clumping is generally a result of unhappy/ unhealthy neurons trying to find partners to "feed" off of, so perhaps the neurons are not finding enough goodies in your growth media to keep them happy.   I use NbActiv4 from Brainbits, LLC, a serum-free complete media.  I replace half the media about 3 days after plating, then once weekly thereafter.  Also, though you don't suspect your substrate, I use only high molecular wt PDL (Sigma P7405) coating for my 12mm glass coverslips.  You might consider trying this.  Good luck, hope this helps!

Hi Cedric, 

Thanks for your kind help. I will contact you if we need it.

Regards,

Youhong

I do not have enough information on the subject.

Thank-you to Ulrich. Very interesting paper by van Goethem. Figure 4 does show some difference between Lister and Long-Evans in with regard to the Memory impairing effects of scopolamine although I would have liked to see the exact same doses on the x-axis. However they are much more similare to each other than they are to Wistars. As Robin said, they are the same species but they are listed as a different rat by charles river and over the years, I have learned that subtle things can matter which is why I asked the question. This would be a very expensive test to do in Canada as I belive the Lister rat is only available in Europe and then you have to account for the stress difference in shipping. Thank-you again

Are you talking about on human patients or laboratory animals? You can always put in an arterial line with pressure transducer hook-up with the plethysmography to monitor BP in real-time. But you do need to know how to calibrate and zero the machine. You can blindly insert the angiocatheter into an artery or do cut-down. Blind insertion is simple but needs a lot of practice.

Don't forget control groups, especially for the more volatile diluents. We usually also include a saline injected group, if that is what your compound is diluted in, just to control for the stress of chronic injections.

Best of luck.

Hi Annadurai

Depending on what the exact need you have for capillary blood is, the analogous sampling methods to a finger prick in rodents is probably the dorsal pedal vein sample (prick the vessel on top of the foot and collect using a capillary tube). You can also do a prick sample from the tail vein and collect using a capillary tube.

The following reviews might help you out as well:

Hope this helps

Pete

Retroorbital is indeed painful and should be avoided whenever possible, since rats are our friends!

...what about the tail vein (especially for small volumes an repeated sampling)? The tail can easily be disinfected to avoid infection.

Hi,

I would suggest using the fine-tipped tweezers (see below for image) and removing this tissue starting at the olfactory bulb. After decapitating the animal and removing the skull, you will see like a black connective tissue around the olfactory bulb. Use the tweezers to break it apart and pull it off (carefully). You should then repeat this on the other side and the brain should be free.

Yes sir..

Surgical or chemical ? Chemical blockade is very efficient and no bleeding.

To stop bleeding use either hypertonic saline or soap foam - prepared by you from solid soap and some water

Many thanks Dr Christopher for these informations

More detail is needed to properly address this question. SD rats are widely used in many fields and easier to handle, however, SDs also have some drawbacks. Endpoints are also critical as many behavioral testing has differential effects across strain, and may or may not correlate to previous literature. Again details are necessary for age and sex. Female rats of any strain have several factors that need to be considered, such as estrous cycle in normal cycling females, OvX females with and without hormone replacement, or post-menopausal females if your study is in the geriatric population.

For retro-orbital bleeds in mice I use long Pasteur pipettes irrespective the size of an animal. A good thing about this approach is that you never fully fill the pipette with the with blood as its capacity often exceeds the amount of animal's blood.