2nd round of mice sacrifice!

Today 7 mice were sacrificed with a rat as per schedule.

The tissues from the mice – testis and ovary are collect in Bouin’s solution and allowed to get fixed.

The tissues which were collected last week, which waited till today, getting ‘fixed’ was given to Department of pathology for further processing, paraffin embedding, sectioning and staining. Though they were unhappy about the extra work they got because of our tissues, I am relieved as I don’t have to struggle in my department putting up the lab. More over I don’t have to slog with our assistant professors for the preparation of histological slides.

Out of 7 mice dissected one female mice was concieved and it had 5 intrauterine concepts.

Some of the images taken during dissection are below.

Abdominal exposure of the mice, intestines are out. Working in the area of kidney and uterus in an attempt to isolate the ovary.

This is me and BK sir. We dissected mice in our dissection hall - in the gap between the practical and viva voce for the 1st year MBBS students, 1st sessional examinations are on!

Pregnant uterus dissected intoto with the ovaries and Fallopian tubes

4 Mice died! But Experiment is ON.

It was all fine for the first two days. On the third day I found a female mice dead from the 4 mg group. I took the corpus and dissected it in our department with SK sir. But by the afternoon, it was badly smelling and we could not bear it till we could possibly find its ovaries! And I have no idea where its ovaries were! I got some fat from the omentum about the calculated size of gonad and assumed it as ovary!!

The next day – while taking dissection I got a call – Akka, informed me that another mouse is dead. It was again a female, from the group which received 4mg drug. I discarded the corpus straight away!

The next day – Sunday – after the morning ‘tulu mass session at Milagris’ we entered the animal house taking the name of the god and found a male (8mg group) and a female (4 mg group) dead. I know it is unscientific in any study to discard the corpus without knowing the cause of death.

Since then there are no more casualties.

Today I and BK sacrificed 5 mice (3 male+ 2 female) by simple cervical dislocation. We could identify the ovaries and pancreas which we had difficulties in earlier attempts. I have taken the tissues – testis and ovaries in Bouin’s solution and pancreas and kidney in regular 10% formalin.

The story so far-  all mice were injected with Gemcitabine on 23-11-10.

No. of mice injected with drug	Approximate amount	Mice dead	Date of death	Number of mice sacrificed to be on
				01-12-10	07-11-10	30-01-11
6 male mice	4 mg each			2	2	2
4 male mice	8 mg each	1	28-11-10	1	2
5 female mice	4 mg each	1 1 1	26-11-10 27-11-10 28-11-10	1	1(-1)	0
4 female mice	8 mg each			1	2	1(+1)

We are left with 19-4-5=10 mice.

Next Tuesday – 7 mice are scheduled to die.

I am left with 3 to test the reversibility of the drug later in January.

The histological processing of the tissues have started, we are running a trial, by using human liver and rat spleen. Tomorrow it will take the proper shape. By then I will process the mice tissues and get ready for the sectioning and staining by Thursday.

Will post the photos of mice tomorrow.

Experiment started!

Today we injected the drug!

Some how the department of Pharmacology is persuaded and they agreed to give the aged mice - which they are not using. Yesterday Prof and Head of Pharmacology directed Akka to give the mice. I received 10 male and 9 female Swiss albino mice in two cages. I made the necessary purchasing of diluent for Inj. Gemita - that's NS, with 20 insulin syringes.
Today - with BK sir and Santhosh sir we injected the mice with Gemcitabine and divided them into 2 groups - those which got 4mg and those which got 8mg!

No. of mice injected with drug	Date	Approximate amount	Number of mice sacrificed to be on
			30-11-10	07-11-10	30-01-11
6 male mice	23-11-10	4 mg each	2	2	2
4 male mice	23-11-10	8 mg each	2	2
5 female mice	23-11-10	4 mg each	2	2	1
4 female mice	23-11-10	8 mg each	2	2

Totally 19 mice are subjected to the study and we have diluted the entire drug in the ampule !!
We are left with more than half of the drug - most probably will be discarded tomorrow !

Tomorrow will be posting the photos !

Mice procurement - not so easy!

The department of Pharmacology has established the central animal house and in charge of it. It has taken a heavy toll on the personnel involved in the work, and are in colossal hope of the rewards for their solid work.

From our side – we always are in tough fight against materialistic attitude of the people around us, making fun of people, are using the same material which they are bargaining for their survival! It’s a tough call, but every time an egoistic face will unearth and will wipe the whole idea of collaboration and corporation.

So in these conflicts the idea of whose animal was it, which invariably not going to make up for the race of survival but eventually going to die – will stand as a mockery of all the human efforts on the surface of earth and eruct the walls of hatred for materialistic, mortal men and at the same time, make us feel the eternity of the concept that we are earthly!

Of late we came to know that the people in pharmacology didn’t like the concept of using the animals by us, which they have purchased for money.

I have made a clear notion- that the animals for our experiment will be purchased just as they are doing. Probably that’s the only way of making it.

I also got the clear notification that BK can get away with it. And any irregularity on the part of animal procurement, handling and sacrificing will have a serious face and consequences if unforeseen mishappening occur with me.



Mice work - Yet to start

For various reasons the mice experiments are getting delayed.

1. Our histology lab is not yet funstional - we have ordered for tissue capsules, it is on its way - from past 2 weeks.
2. The rats have become intolerant in BK's experiment - it can no more be fed like the previous one. Thay are totally restless and agitated during every attempt of feedinf oflox. New methods are on cards. Have to implement.
3. Pathology people are more cooperating with us in seeding a lab establishment than in processing the tissue given to them. We are in the mood of having our lab functioning - than to collaberate with them!
4. Of late - a cold war has started - with us and Pharmacology department - sparked by uninformed use of one more rat for the oflox experiment. We have given lengthy request letter for more rats and mice. Finally they made the comment, much inevitable - 'we dont have enough male mice, you may have to order'. This may have more impact in future than it is evident today!

From today we have started writing in a movememt kind of register in the central animal house, indicating the time of visit, the purpose of visit, the persons visited to the central animal house.
Some how we have to see that some staff member is present whenever we goto animal house, so that any mishappenings can be overcome easily! and my senses are telling that the animal house is not all that fool-proof.

Histology of rat callus formation

Histology of mice testis

These are the photographs of histology of normal mice testis

Schedule

This is the proposed schedule for the mice experiments:

No of mice injected with drug	Date	Approximate amount	Number of mice sacrificed on
			15-11-10	22-11-10	08-01-11
3 male mice	08-11-10	3-4 mg each	1	1	1
2 male mice	08-11-10	6-8 mg each	1	1
2 female mice	08-11-10	3-4 mg each	1	1

Totally 7 mice will be subjected to the study and I need 40 mg of the drug.
I am delaying it as I am going home on next week for Diwali

Got Gemita!

It was just like going to any shop buying an article. BK droped both of us, me and Santhosh in Bendur, near Dominos pizza shop. As was said Mediglobe, located just at its back. We walked in introduced myself, handed over my synopsis copy and within 2 minutes they put a small ampule in the plastic cover and handed over to me. We walked out. Some Rs 1400 saved.!

I will start the experiment soon..
probably tomorrow.

Drug procurement

Procurement

Drug required is Gemcitabine – 200 mg inject able.

I searched in the CIMS online and found 14 different brands of it with prices varying from Rs.1050 to Rs.1920. My intension is to go for the cheapest possible brand, obviously.

Long time back Sudha has promised me of getting it for free as she is in pharmacology, and as she can get to know the men at work selling this by some means and fetch it for free. She even called me when she was just about to enter the room of Dr. Krishna Prasad, the medical oncologist, for getting the drug called Dasatinib, for her presentation of a paper in national conference this December. I had denied the offer.

From the past week I have decided to start the experiment. The first part is over as the control mice dissection is done. I went to AJ pharmacy this Sunday, enquired about the drug, which they in deed had, but the costliest one. Its name is Gemicite. It would cost me Rs.1920. But the pharmacy manager told me he can order the other lower cost brands.

It was BK sir who told me to give a try in Gopal Kamath wholesale drug distributors as they are the largest drug marketers in Mangalore, supplying most part of the drugs prescribed here.

I went to the shop Monday, the day I was fooled by wrongly read news in Hindu about the 9^th chapter discloser by Swami Brahmananda in TMA Pai conference hall. They had the drug. It costs Rs. 1119. I didn’t have the money then. I decided to buy on the following day.

The next day I and Santhosh sir walked to the shop and got the bill for the 200mg drug paid the amount. Including the owner of the shop searched the whole godown and finally announced that the drug is not present with them. They gave the number of the Dabur pharma medical representative, Mr Ravikumar.

I spoke with him and as instigated by Santhosh sir, I asked the drug for free. He said they can give it – if I submit the letter by an oncologist stating that the drug is for the study or for poor patient. I agreed for the letter submission.

Later I found that the only visiting oncologist to AJ is Dr. Krishna Prasad and it is very hard to locate him. He comes for the rounds at 6am if he had patient(s).

When I said the story to BK, he said don’t get the drug for free and tell the medical rep that I pay for it for the reasons we cant go behind an oncologist (just) for the letter.

I called him and said I can give you my thesis synopsis and letter of registration with RGUHS. He agreed to give the drug by 2 noon!

By 5.30 pm today he called and asked me to collect the drug from some shop in Dambale.

I will go and get the drug tomorrow!

I found mice - dead!

Every day I and BK sir use to visit the animal house at least once, for giving oflox to the rat. After one set of rats were sacrificed, the new ones are fractured, drug is started in a week’s time. We failed to visit the animal house last Monday.

And that day- 4 of the mice were dead, for the reasons unknown. We got the news on the following say. I gave my personal number to the ‘akka’ incharge of the animal house and asked her to inform the news of any animal death in there irrespective of cause of death. She understood that people from anatomy department are after dead – anything dead!

This Monday I got the call at 8 am, akka said, “ 2 mice are dead”. I was relieved that I don’t have to kill normal mice on my own to run my control study.

I was getting ready for the dissection – found that Ajay too interested in the mice dissection. BK sir joined. We dissected both the mice. We took out the testis, pancreas and liver. Sperm extraction from the epidydimis was not possible as it was already shrivelled.testis is put in Bouins solution and rest organs in the formalin. It’s still with us now. Tomorrow will hand it over to pathology for the histological study.

This will be my first mice tissue sample collection.

Today I am going to get the drug Gemvita, from Gopal Kamath wholesale distributors. It will cost me Rs. 1150 for 200 mg amp. It can be stored at room temperature. But the problem is it said it has to be used within 24 hrs of reconstitution.

In our animal house there is no proper ventilation and temperature maintenance. This could be the probable reason for the death of the mice, as no one attends them on whole of Sunday. They are not given water and food, and not even fresh air on Sundays. A strict holiday. In 2 consecutive weekends we have lost 6 mice.  

Oflox experiment

It all started with BK sir's experiment of oflox on fracture healing.

We had fractured femurs (femora) of two 300 gm wistar rats on 19/09/2010, taken to Department of radiology - took X ray of the fractures and started them on oflox BD doses a week after. Rats recieved ofloxacin for 3 weeks.

On 20/10/2010 we killed  both the rats using KCl, dissected them and taken out the femur with callus. Along with that we have also taken other tissues - testis, liver, pancreas and normal knee joint.

We gave all the tissues for the histopathological examination to Department of Pathology. The results are awaited.