Pyruvate Tolerance Test - Revision history

Davebrid: added context to PTT

2022-04-06T17:50:18Z

added context to PTT

← Older revision		Revision as of 17:50, 6 April 2022
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			Pyruvate tolerance tests are typically done to understand the rates of pyruvate conversion into glucose (as an indirect measure of gluconeogenesis). The assay is relatively straightforward, we fast mice for ~6h to normalize glycemia then perform intraperitoneal injections of pyruvate. That pyruvate is converted into glucose, primarily by the liver. The rate of glucose production from pyruvate is indicated by the peak, or area under the curve of the glucose-time curve. A couple of key caveats. Several substrates could be used for this including pyruvate, glycerol, lactate, alanine, or glutamine, but pyruvate appears to be the most common in the literature. Second, you should '''always''' perform a [[Glucose Tolerance Test\|glucose tolerance test]] to interpret these results. This is because if you have impaired glucose disposal, you will also have elevated disposal of pyruvate-elicited glucose. That would mean that glucose intolerance could be interpreted as excessive pyruvate-glucose conversion. While this assay is relatively easy and inexpensive, the gold standard of glucose production ''in vivo'' is to monitor glucose production during a euglycemic clamp either in response to insulin (suppressive to gluconeogenesis) or glucagon (promoting it).

	==Materials==		==Materials==
	* Glucometer - AccuChek Advantage		* Glucometer - AccuChek Advantage
	* Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]		* Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]
	* Scale		* Scale or echoMRI to determine mouse mass or mouse lean mass
	* Beaker for weighing mice		* Beaker for weighing mice
	* Syringes		* Syringes
	* 2.5g pyruvate in 10 mL PBS- This will correspond to 2.5 g/kg injections.		* 2.5g pyruvate in 10 mL PBS, dissolve then adjust pH to 7.3-7.5 - This will correspond to 2.5 g/kg injections.
	* Timer		* Timer

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	==Protocol==		==Protocol==
	* Remove food from mice for about 6h.		* Remove food from mice for about 6h.
	* Typically starve the mice at 9AM and aim to start injections at ~~3 PM~~		* Typically starve the mice at 9AM and aim to start injections at 3PM
	* Weigh mice, mark tails if necessary and take fasting glucose measurement via a tail clip.		* Weigh mice, mark tails if necessary, and take a fasting glucose measurement via a tail clip.
	* Prepare pyruvate syringes with 10 uL per g mouse ~~weight~~ (ie for a 30g mouse, 300 uL).		* Prepare pyruvate syringes with 10 uL per g mouse mass (''i.e.'' for a 30g mouse, 300 uL). Ideally, injections are per lean mass, not total mass if the animals differ substantially in adiposity. This is since higher levels of adiposity would result in larger pyruvate injections and potentially inaccurate higher glucose responses.
	* At approximately 1 min intervals, inject appropriate amount of pyruvate into ~~interperitoneal~~ cavity of the mouse.		* At approximately 1 min intervals, inject appropriate amount of pyruvate into intraperitoneal cavity of the mouse.
	* Immobilize mouse and restrain tail with one hand		* Immobilize mouse and restrain tail with one hand
	* Aim needle between the midline and the hip bone		* Aim needle between the midline and the hip bone
	* Insert syringe (do not inject) into cavity		* Insert syringe (do not inject) into the cavity
	* Eject syringe.		* Eject syringe.
	* At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.		* At desired intervals (normally 15, 30, 45, 60, 75, 90, 105, and 120 min), take blood glucose measurements from the tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.
	* Analyse data by both % change from fasting glucose and absolute values.		* Analyse data by both % change from fasting glucose and absolute values. You may also want to calculate the initial rate of glucose increase (see [[Rate of drop analysis\|this protocol]] for details), or the area under the glucose-time curve.

Davebrid: fixed typo

2017-04-27T16:39:23Z

fixed typo

← Older revision		Revision as of 16:39, 27 April 2017
Line 12:		Line 12:
	* Remove food from mice for about 6h.		* Remove food from mice for about 6h.
	* Typically starve the mice at 9AM and aim to start injections at 3 PM		* Typically starve the mice at 9AM and aim to start injections at 3 PM
	* Weigh mice, mark tails if necessary and [[take fasting glucose measurement]] via a tail clip.		* Weigh mice, mark tails if necessary and take fasting glucose measurement via a tail clip.
	* Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).		* Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).
	* At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.		* At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.

Davebrid: Updated layout of the protocol

2017-04-27T16:35:10Z

Updated layout of the protocol

@@ Line 1: / Line 1: @@
-'''Materials'''
+==Materials==
-Glucometer - AccuChek Advantage
+==Protocol==
+* Remove food from mice for about 6h.
-Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]
+* Typically starve the mice at 9AM and aim to start injections at 3 PM
+* Weigh mice, mark tails if necessary and [[take fasting glucose measurement]] via a tail clip.
-Scale
+* Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).
+* At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.
-Beaker for weighing mice
+* Immobilize mouse and restrain tail with one hand
+* Aim needle between the midline and the hip bone
-Syringes
+* Insert syringe (do not inject) into cavity
+* Eject syringe.
-.5g pyruvate in 10 mL PBS- This will correspond to 2.5 g/kg injections.
+* At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.
+* Analyse data by both % change from fasting glucose and absolute values.

Iharvey at 19:40, 23 June 2015

2015-06-23T19:40:47Z

← Older revision		Revision as of 19:40, 23 June 2015
Line 1:		Line 1:
	'''Materials'''		'''Materials'''


	Glucometer - AccuChek Advantage		Glucometer - AccuChek Advantage

	Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]		Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]

	Scale		Scale

	Beaker for weighing mice		Beaker for weighing mice

	Syringes		Syringes

	2.5g pyruvate in 10 mL PBS- This will correspond to 2.5 g/kg injections.		2.5g pyruvate in 10 mL PBS- This will correspond to 2.5 g/kg injections.

	Timer		Timer


	'''Protocol'''		'''Protocol'''


	Remove food from mice for about 6h.		Remove food from mice for about 6h.

	Typically starve the mice at 9AM and aim to start injections at 3 PM		Typically starve the mice at 9AM and aim to start injections at 3 PM

	Weigh mice, mark tails if necessary and [[take fasting glucose measurement]] via a tail clip.		Weigh mice, mark tails if necessary and [[take fasting glucose measurement]] via a tail clip.

	Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).		Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).

	At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.		At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.

	Immobilize mouse and restrain tail with one hand		Immobilize mouse and restrain tail with one hand

	Aim needle between the midline and the hip bone		Aim needle between the midline and the hip bone

	Insert syringe (do not inject) into cavity		Insert syringe (do not inject) into cavity

	Eject syringe.		Eject syringe.

	At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.		At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.

	Analyse data by both % change from fasting glucose and absolute values.		Analyse data by both % change from fasting glucose and absolute values.

Iharvey: Created page with "'''Materials''' Glucometer - AccuChek Advantage Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2..."

2015-06-23T19:38:46Z

Created page with "'''Materials''' Glucometer - AccuChek Advantage Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2..."

New page

'''Materials'''

Glucometer - AccuChek Advantage
Glucose Test Strips - AccuChek Comfort Curve. Order these through Materiels Service by calling 936-6077 and ordering product 2535. For information see [here]
Scale
Beaker for weighing mice
Syringes
2.5g pyruvate in 10 mL PBS- This will correspond to 2.5 g/kg injections.
Timer

'''Protocol'''

Remove food from mice for about 6h.
Typically starve the mice at 9AM and aim to start injections at 3 PM
Weigh mice, mark tails if necessary and [[take fasting glucose measurement]] via a tail clip.
Prepare pyruvate syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).
At approximately 1 min intervals, inject appropriate amount of pyruvate into interperitoneal cavity of the mouse.
Immobilize mouse and restrain tail with one hand
Aim needle between the midline and the hip bone
Insert syringe (do not inject) into cavity
Eject syringe.
At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.
Analyse data by both % change from fasting glucose and absolute values.