Lab 10: Insulin and Glucose Tolerance
Objectives:
1) Understand the role of insulin and glucagon secretion for the regulation of plasma glucose homeostasis
2) Understand the differences between Type 1, Type 2 and Gestational Diabetes and how a glucose tolerance test can be used to detect these conditions.
3) Understand the differences and physiological significance of euglycemia, hypoglycemia and hyperglycemia, the units of plasma glucose measurement and the ranges of values associated with these conditions.
4) Understand postprandial glucose uptake and distribution into the blood stream.
5) Understand how to interpret blood glucose made using a finger blood sample and glucose analyzer.
Background:
The human body needs a constant supply of ATP to carry out just about any essential cellular process (i.e., active transport, muscle cell contraction, transmission of neural signals, etc…). While the body can utilize fatty acids and some amino acids to generate ATP through aerobic respiration, most of the body’s ATP generation comes from the metabolism of carbohydrates (i.e., glucose). In fact, some tissues (nervous system) rely almost exclusively on ATP generated from glucose metabolism. As a result, it is absolutely critical that the body maintain adequate levels of glucose in the bloodstream and prevent blood glucose levels from dropping too low, a condition known as HYPOGLYCEMIA; below 70 mg glucose/dl). However, long term, chronic elevation of blood glucose levels (HYPERGLYCEMIA; above 140 mg/dl) also is detrimental to the body. Uncontrolled high blood glucose levels are known to result in damage to the retina of the eye (retinopathy), neurons (neuropathy), and smaller blood vessels (especially in the kidneys!). These are just a few of the detrimental effects of uncontrolled Type 1 and Type 2 diabetes mellitus.
The human body maintains normal (euglycemic) blood glucose levels somewhere in the range of 70-120 mg/dL. These numbers will vary from one person to the next and will certainly fluctuate throughout the day as we go through periods of fasting and feeding. An early morning glucose level after not eating for 8-10 hours will commonly be near the lower range (70-90 mg/dL) while blood glucose levels will spike following a meal or snack (often 150-190 mg/dL).
The body employs several hormones to limit hyperglycemia or hypoglycemia. When our glucose levels drop in between meals or snacks, the pancreas releases GLUCAGON which promotes the conversion of glycogen polymers in the liver and skeletal muscle into glucose monomers which are released into the bloodstream which elevates our blood glucose levels. This is known as GLYCOGENOLYSIS. Glucocorticoids such as cortisol also play a more restricted role in regulating blood sugar levels during times of stress. Under these conditions, cortisol promotes the generation of glucose from non-carbohydrate sources such as some amino acids. This process is known as GLUCONEOGENESIS. Both of these processes function to increase our blood glucose levels. After a meal, our blood glucose levels rise dramatically triggering the release of INSULIN from the pancreas. Insulin promotes the uptake of excess glucose from the bloodstream and eventually stimulates its conversion to glycogen for storage. In VERY SIMPLE terms, one can think of insulin and glucagon carrying out similar processes, but in opposite directions. Glucagon promotes the hydrolysis of glycogen into glucose (i.e., glycogenolysis) leading the increased blood glucose levels during fasting. Insulin promotes the opposite (i.e., leading to decreased blood sugar levels).
A person’s ability to regulate blood glucose levels is not only dependent on the proper release of these hormones, but also on their ability to influence the functions of target cells in the body. As we know, Type 1 diabetics have lost the ability to produce and secrete insulin when needed which prevents the body’s cells and tissues from absorbing and utilizing the glucose in the bloodstream. The causes of Type 2 diabetes are a bit more complex but often are related not to low insulin levels per se, but to the inability of the body’s cells to respond to insulin due to alterations in the insulin receptor or a malfunction of second messenger systems after insulin binds to its receptor. Gestational diabetes occurs in women during pregnancy and usually resolves after pregnancy is complete, although the infant takes on a higher risk for diabetes, obesity and heart disease as an adult as a result of gestational exposure to excess glucose. Individuals are often referred to as being “insulin resistant” because they may be able to secrete insulin from the beta-cells of their pancreas, but the insulin is unable to cause insulin sensitive cells to become permeable to glucose, hence their plasma glucose concentration does not rapidly return to baseline postprandially.
.
An important clinical test known as a GLUCOSE TOLERANCE TEST (GTT) is performed on individuals that are suspected to be suffering from diabetes. The goal of the test is to see how well your body responds to a sharp increase in blood glucose levels, otherwise known as a “glucose challenge”. It is important to note that this test does not by itself determine the type of diabetes a person might have, but rather tells the physician that the person likely has some type of problem regulating their blood glucose levels. Briefly, a person undergoing a GTT will fast for approximately 8 hours in order to get their blood glucose levels down towards the lower end of the normal range. The person then drinks a sugary beverage that contains about 75g of glucose, roughly the amount of sugar in a 20 oz. bottle of regular Coke or Mt. Dew. The glycemic challenge used in this laboratory activity will be a single 12 oz can of lemon lime Sierra Mist (37 gram carbohydrate challenge), which will be compared to Diet Sierra Mist (flavor but no sugar) or water (no flavor and no sugar). For Nutrition Information see Table 1.
The person’s blood glucose is measured BEFORE drinking the beverage to determine a baseline glucose level and then every 20-30 minutes afterwards to see how the body responds to the glucose challenge. Blood glucose levels will typically peak about 45 minutes after drinking the beverage and then (hopefully) decline fairly rapidly thereafter. A healthy individual will see their blood glucose levels jump to 130- 180 mg/dL after drinking the beverage and then see a steady decline over the next 90 minutes or so back down to normal levels. A person with diabetes or who is insulin resistant will also see their glucose levels jump up but those levels will stay elevated for a much longer period of time (often hours). Basically, this is the test that we will be doing in lab.
An important additional test that is often done is to measure the levels of insulin in the bloodstream along with the glucose levels to help differentiate whether a person suffers from gestational, Type 1 diabetes or Type 2 diabetes.
After consuming the sugary beverage the blood glucose levels will rise and the pancreas responds by increasing the secretion of insulin to help restore normal blood glucose levels more rapidly. A sharp increase in insulin secretion might be seen in a person with Type 2 diabetes (insulin resistant) but not in a person with Type 1 diabetes since their pancreas has lost the ability to produce and secrete insulin. In essence, an increase in insulin secretion without a corresponding drop in blood glucose levels suggests that the person’s cells or tissues are not responding to the insulin properly (i.e., they are insulin resistant).
Table 1. Nutrition Facts for the three test beverages used in lab.Lemon Lime Sierra Mist (12 oz can) / Diet Sierra Mist (12 oz can) artificially sweetened with Aspartame and Acesulfame K
12 oz glass of water: 355 ml 0 calories
Protocol for Experiment:
1. Form groups of 2-5 students based around a volunteer who wishes to consume the test beverage (water, Diet Sierra Mist or Sierra Mist). This one person per group will consume the test beverage and they will have 4 finger-stick blood glucose tests. All beverages must be consumed in hallway, for purposes of safety no beverages can come into the lab.
2. Subject who drinks beverage should have ideally fasted overnight, but a 6 hour fast prior to the beginning of the test will be fine for this lab activity. No coffee, soda, pop-tarts, etc. Water is definitely OK though. At any rate keep track of the time of the last meal for the volunteer.
3. A “fasting blood glucose” should be measured BEFORE drinking your beverage. Your lab instructor will instruct you on how to measure blood glucose using the blood glucose meters in lab.
a. Briefly, you will need (1) alcohol wipe, (1) lancet, (1) glucose test strip, (1) glucose meter for each measurement.
b. Insert a fresh, unused glucose strip into the glucose meter
c. Wipe the end of your finger with the alcohol wipe
d. Use the lancet to produce a small drop of blood
e. Place end of the glucose strip next to the blood drop so that it is drawn up into the end of the strip.
f. Record your glucose reading
4. Drink your assigned beverage as quickly as possible (do not take more than 2-3 minutes to do this). TAKE NOTE OF THE TIME YOU FINISHED DRINKING YOUR BEVERAGE
5. After 20, 40 and 80 minutes, re-measure your blood sugar and record the reading on the data sheet provided, but only drink the beverage once at the start of the study (after baseline blood glucose measurement).
Blood Sample Collection:
Put on gloves and keep all materials at the station next to the window. At the station you will use a Unistik to draw blood from the finger. Place all contaminated materials in red biohazard bags!
1) Wipe lateral edge of finger with alcohol swab.
2) Let it dry before drawing blood (it hurts if alcohol has not dried off)
3) Follow instructions in illustrations below:
PLACE UNISTIK FIRMLY AGAINST FINGER PRIOR TO RELEASING THE FIRE BUTTON, it will make a click sound and the lancet will strike then retract.
4) Immediately throw Unistik in “Sharps Container” after use… DO NOT REUSE!
5) Put finger below level of heart and let a drop of blood form, you may “GENTLY” milk blood out of the finger if a drop of blood does nor readily form.
LAB WRITE UP: (Hand in to your Lecture Professor on Monday, April 27, 2015)
Please answer the following questions and them in along with your Group Lab Report to your lecture instructor. Answers must be typed, grammatically correct and in complete paragraphs! Answer as items #1, #2a, #2b, #2c and #3 and complete lab sheet.
1. Based on your data how does consuming a sugar-containing beverage, an artificially sweetened beverage or water modify ones blood glucose? How or why did you come to this conclusion?
2. Based on the data that was collected discuss what you think was occurring over the course of the experiment.
a. Specifically discuss the response of the pancreas (i.e., secretion of BOTH insulin and glucagon) as well as the effects of those hormones on tissues in the body.
b. Do you think insulin secretion increased or decreased? Do you think glucagon secretion increased or decreased?
c. How might the timing of your last meal/snack/beverage have influenced your glycemic response?
3. Frequently, additional symptoms of uncontrolled diabetes are (1) GLUCOSURIA, (2) POLYURIA (frequent urination), and (3) POLYDIPSIA (excessive thirst). Based on our knowledge of the urinary system, fluid homeostasis, and the fact that glucose contributes to the overall osmolarity of tissue fluids (including tubular fluids) explain why prolonged hyperglycemia can cause these THREE symptoms.
DATA RECORD for Lab Group and group lab report
Group Members: ______
______
______
______
DO NOT IDENTIFY YOUR VOLUNTEER IN YOUR GROUP LAB REPORT
Day of Lab Attendance: ______
BEVERAGE CONSUMED ______
TIME BEVERAGE WAS CONSUMED ______
Time Taken / Glucose Reading (mg/dL)Baseline
20 minutes
40 minutes
80 minutes
What time did you last consume a meal, snack, or caloric beverage? ______
What was the meal, snack, or caloric beverage? ______
______
______
______
Notes: ______
______
______
______
Plot your Glucose results on the following graph. Then compare your results with groups that drank an alternative substance.
COPY OF DATA RECORD for Lab Instructor
Group Members: ______
______
______
______
Do not identify your volunteer
Day of Lab Attendance: ______
BEVERAGE CONSUMED ______
TIME BEVERAGE WAS CONSUMED______
Data from all subjects and all treatments will summarized and presented to each lecture. You can compare average treatment responses for all three groups and discuss why responses might vary.
Time Taken / Glucose Reading (mg/dL)Baseline
20 minutes
40 minutes
80 minutes