Acute stress responses in salivaryalpha-amylase predict increases of plasma

norepinephrine

Myriam V. Thoma, Clemens Kirschbaum, Jutta M. Wolf, Nicolas Rohleder

Journal from Biological Psychology volume 91 (2012) page 342-348

Present by Kamonwan Khotchapan

OutlineIntroduction

Psychosocial stress Salivary glands Production of saliva Catecholamine Autonomic nervous system Salivary and norepinephine

ObjectiveMaterials and Methods

Procedures for Trier social stress test (TSST)Biochemical analysesPsychometric measurementsStatistical analyses

ResultsDiscussion and conclusion

Psychosocial stress

Psychological stress occurs when an individual perceives that

environmental demands tax or exceed his or her adaptive

capacity and it affected in sympathetic nervous system (SNS) activity

When psychosocial stress triggers a stress response, the body releases a group of stress hormones including cortisol, epinephrine (or adrenalin) and dopamine which lead to a burst of energy as well as other changes in the body

Psychosocial stress

Psychosocial stress was determined by measuring catecholamine in blood plasma or using electrophysiological methods

- Skin conductance

- Heart rateHowever, these methods are complicated and invasive methods,

therefore, using non-invasive method was interested to determine

psychosocial stress such as investigated salivary alpha-amylase

Saliva glands

http://reasonablywell-julia.blogspot.com/2011/05/facial-swelling-and-sjogrens-syndrome.html

Production Of Saliva

http://www.zuniv.net/physiology/book/chapter22.html

Sympathetic cAMP secreted proteins

Parasympathetic Phospholipase C Ca2+ Secreted

fluid

Salivary alpha-amylase(sAA)

http://chemistry.tutorvista.com/organic-chemistry/glycosidic-linkage.html

sAA (α-1,4-α-D-glucan 4-glucanohydrolase) It is a digestive enzyme that hydrolyses alpha-1,4 bonds of large polysaccharides such as starch and glycogen, yielding the smaller by-products of glucose and maltose.It consists of two families of isoenzymes

• glycosylated• no carbohydrate

It accounts for 40 – 50 % of total salivary gland-produced protein

catecholamine

norepinephrine

epinephrine

dopamine

A catecholamine (CA) is a monoamine.

CA are compose of …• norepinephrine• epinephrine• dopamine

Source

• adrenal medulla

Effects• physiological changes (fight-or-flight response). • increases in heart rate, blood pressure, blood glucose levels• a general reaction of the sympathetic nervous system

Autonomic nervous system

http://www.khironhouse.com/blog/a-key-relationship-for-trauma-treatment-the-sympathetic-and-parasympathetic-branches-of-the-nervous-system/

Why do research determine salivary alpha-amylase and norepinephine

http://www.hakeem-sy.com/main/node/16265

Norepinephrine from sympathetic neurons binds to bothalpha- and beta-adrenergic receptors on the acinar cell and more sensitivity with β2 receptor .

Acute response of salivary -amylase and plasma norepinephine to the laboratorystress paradism TSST in n = 12 healthy men and women

Rohleder et al., Ann. N.Y. Acad. Sci. 2004; 1032: 25-263.

In previous study

Trier social stress test (TSST)

Objective

• To examine the correlation between stress-induced sAA activity with NE and E .

Materials and Methods

Subject

40

18 use oral contraceptive(6 smoke)

22 in luteal phase

26 (9 smoke)

BMI 17-27 kg/m2

Age 18-24 yearsFree of chronic stress Free of medication (Ex. Beta-block ,glucocorticoid)

free of chronic disease (Ex. Endocrine, cardiovascular)

Procedures

1

• At first use salivettes to keep a saliva and keep blood sample to EDTA tube.

2• All subject do the ‘Trier social stress test (TSST)’

3• After TSST a second blood draw and saliva sample

was taken again.

4

• At 20 min and 40 min taken a blood sample and saliva sample again.

Trier social stress test (TSST)

For TSST has divided into 2 room

Room A

Room B

Biochemical analyses

Use enzyme kinetic method was processed on Genesis RSP8/150 liquid handing system

Dilute sample at 1:625 and use 20 µl of dilute saliva and standard transferred into 96 well microplate

Standard prepared from calibrator solution with concentration 326, 163, 81.5 , 40.7, 20.38 ,10.19 and 5.01 U/L of alpha amylase

1. Salivary alpha- amylase (sAA)

Procedures

Pipette 80 µl of substrate reagent to each well

Incubate at 37 ºc in water bath 90 s

Measurement at 405 nm

Incubate at 37 ºc again for 5 min

Measurement at 405 nm

water bath

Biochemical analyses

2.Norepinephrine and Epinephrine

Plasma sample was analyze for NE and E by

high-pressure liquid chromatography (HPLC) at the Laboratory for Stress Monitoring

Psychometric measurements

Perceived stress

Depressive symptoms

Psychometric measurements

Perceived stress

Its was assessed with the 14-item version Perceived Stress Scale (PSS) . PSS is measure the level of stress by an individual experience by ranging from 0 ‘never’ to 4 ‘very often’

Psychometric measurements

Depressive symptoms

This symptom were assessed with the 20-item

of the Center of Epidemiologic Studies Depression Scale (CES-D) that is a self-report questionnaire about the incidence and duration of impairment experienced by individuals depressive affects.

Statistical analyses

Use log 10 test for normal distribution

T-test

x ²-test

For the assessment of difference between group

Results

• Men had a significantly higher BMI than both groups of

women .

• Smokers and non-smokers differed with a trend in terms of sex distribution .

• Results showed no association of depression scale values with stress responses of sAA, norepinephrine, or epinephrine .

Preliminary data analysis

• Perceive stress scale overall mean indicating no elevated perceived stress levels in our sample.

• Results showed no association between PSS values and stress responses of sAA, norepinephrine, or epinephrine .

Preliminary data analysis

Results

Results

Fig. 1. Salivary alpha-amylase (sAA) in response to the acute stress induction

The sAA concentrationsover the course of the fourmeasurement points, we found asignificant change in the sAArelease over time during thestress intervention and highestafter TSST .

ResultsAfter TSST the results shown that the epinephrine responseto acute stress was a weak predictor whereas norepinephinewas higher than epinephine.

Fig. 2. Norepinephrine (NE) and epinephrine (E) in response to the acute stress

Fig. 3. Correlation of stress-induced increases of salivary alpha-amylase (sAA) and norepinephrine (NE).

Results

When correlate between salivary alpha-amylase and norepinephrineresponse to stress we found that p = 0.025 and its significant in psychology .

Correlation of stress-induced increases of sAA and NE

Rohleder et al., Ann. N.Y. Acad. Sci. 2004; 1032: 25-263.

Result from previous study

The results show that the increasing of sAA and NE concentration are correlate in linear regression .

Discussion

The results found significant association between stress-

induced sAA and NE responses, but not between sAAand E responses.

The active secretion of the adrenal medulla contains approximately 80 % E and 20 % NE ; but this proportion is reversed in the sympathetic nerves, which contain predominantly NE , that why NE responses to stress-induced sAA more than E responses.

In the present study we found a significant association between sAA and NE and the usefulness of sAA to function as an indicator for SNS activation.

In addition, we found that stress-induced sAA and Elevels in habitual smokers, in individuals with a lower BMI, and in females using oral contraceptive.

Discussion

From this study show an association between stressresponses of sAA and plasma norepinephrine . Thisrelationship is stronger than the association ofnorepinephrine and epinephrine responses, thus placingthe predictive power of sAA well within the expectedrange for different SNS markers.

Conclusion

Acknowledgements

Dr. Apilak Worachartcheewan

Autonomic nervous system

The autonomic nervous system has two main divisions

• Sympathetic

Promotes a "fight or flight" response, corresponds with arousal and energy generation, and inhibits digestion

• Parasympathetic

Promotes a "rest and digest" response, promotes calming of the nerves return to regular function, and enhances digestion

How to use salivette