Lab 2 (Everted Sac)

Question 1

What is the purpose of the first experiment?

Answer 1

This is to employ an everted sac preparation of the mouse duodenum to investigate the process of Na+ dependent glucose cotransport

Question 2

What are the four protocols?

Answer 2

1. control: Na+ Ringer’s solution
2. Na+-free Ringer’s solution
3. Na+ Ringer’s solution + mucosal phloridzin
4. Na+ Ringer’s solution + serial ouabain

all of these are bubbled with O2

Question 3

What does Phloridzin do?

Answer 3

It blocks the SGLT1 glucose cotransporter

Question 4

What does Ouabain do?

Answer 4

It blocks the Na+/K+ ATPase

Question 5

Which carbohydrate is present in breastmilk? Which two sugars result from the breakdown of this carbohydrate during digestion?

Answer 5

Lactose

Question 6

Baby Jane has diarrhoea. What would this imply about the normal function of her small intestine in relation to water absorption?

Answer 6

It is not absorbing water so she has runny stools. If she didn’t have a functional SGLT1, there would be no osmotic gradient and therefore she would not absorb all of the water.

Question 7

How does Baby Jane’s lack of water absorption relate to her impaired glucose absorption?

Answer 7

Glucose absorption is impaired which affects her osmotic gradient (as Na+ is not being absorbed)

Question 8

Why is Baby Jane dehydrated?

Answer 8

She can not absorb glucose due to a mutation in the SGLT1 glucose cotransporter. This means that there is no Na+ gradient and so there is no osmotic gradient and so she is not absorbing water.

Question 9

How do we know that it is the SGLT1 not the SGLT2 or GLUT2 transporter that is affected in baby Jane?

Answer 9

If we are only looking at the intestine, there could be a problem with the GLUT2 transporter but because she has glucose in her urine, we need to look at the kidney.
SGLT2 is the main transporter to absorb glucose in the kidney. It absorbs 90% of the glucose in the PCT. If she has lots of glucose in her urine then there would be a problem with SGLT2 but because there is only a bit, we know that there is a problem with SGLT1

Question 10

Does Jane have secretory of absorptive diarrhoea?

Answer 10

She has absorptive diarrhoea because she is not absorbing enough Na+ or glucose to absorb water (secretory diarrhoea is due to a problem with CFTR)

Question 11

For the Na+ Ringer’s control sacs, what was the concentration of glucose inside the sac (serosal) compared with the concentration of the mucosal solution?

Answer 11

There was a much higher [glucose] on the basolateral than luminal side (glucose flows from luminal to basolateral)

Question 12

Would the glucose concentration continue to increase inside the Na+ Ringer’s control sac, if you
left the sac incubating for a longer period of time? Would there be a maximum concentration that
could be achieved, or would the concentration continue to increase as long as you incubated the
sac?

Answer 12

The concentration in the sac gets to a point from inside that means we get saturation of both the SGLT1 and GLUT2

Question 13

What was the effect of Na+ free Ringer’s on serosal glucose concentration?

Answer 13

There was a similar concentration on both sides because no glucose could get into the cell as there was no Na+ and SGLT1 requires Na+

Question 14

What was the effect of phloridzin on serosal glucose concentration?

Answer 14

This blocks the SGLT1 but doesn’t bind permanently. This means that only some are inhibited which means that you can still get some glucose can be absorbed and you get some to the basolateral side

Question 15

What is the action of ouabain?

Answer 15

This permanently inhibits the Na+/K+ ATPase so Na+ can’t leave so there is no driver to get Na+ or glucose into the cell via SGLT1