A Practical Approach, Second Edition=Ronald D. Ho.pdf

106 DEVELOPMENTAL REPRODUCTIVE TOXICOLOGY: A PRACTICAL APPROACH, SECOND EDITIONThe small size of some species (e.g., mice) can preclude the collection of large amounts ofblood, limiting the number of end or time points that can be evaluated in a given sample. Investigatorsshould also be aware that there is a marked circadian pattern in the levels of blood glucocorticoids.149 In the rodent, the nadir (~1 g/dl) occurs during the lighted period, with a peak (~20g/dl) during the dark period. Thus, measuring blood glucocorticoids at a single time point can leadto difficulties in interpretation (see Kopf-Maier 135 and Hansen et al. 133 for a discussion of theseissues). There are also multiple reports of strain differences in the responsiveness of the HPA axisin many species. 150–154 Because of the many factors that affect glucocorticoid secretion, investigatorsshould not begin using glucocorticoids as markers for stress until they determine the basal circadianpattern of secretion obtained in their animal facility with the strains they normally use to evaluatechemicals for developmentally toxic effects. Finally, the investigator should be aware that in thepregnant mouse, as well as other species (e.g., rabbit, human), the total glucocorticoid level foundin blood increases dramatically at about gestational day 9, with the peak level observed betweendays 12 and 15. 133,155,156 The day and degree of peak elevation are strain dependent and may berelated to pregnancy-induced increases in the glucocorticoid binding protein in blood that occur ina number of species. 157–1592. CatecholaminesActivation of the HPA axis can also result in the release of catecholamines from the adrenal medulla,as well as from sympathetic nerve endings. Plasma norepinephrine is released primarily fromsympathetic nerve terminals, while epinephrine is secreted exclusively from the adrenal gland. 160These compounds have hemodynamic properties and can alter blood flow to the uterus. Exogenoustreatment of the dam with these substances, as well as other vasoactive agents, such as vasopressin,can cause developmental abnormalities (e.g., Chernoff and Grabowski 161 ). Thus, there has beensome interest in determining their role in maternal toxicity and stress in experimental animals aswell as in humans. 23,137 Cook and colleagues did not observe consistent alterations in blood oruterine catecholamine levels in response to noise, and the levels induced were much lower thanwould be obtained by exogenous treatment with these agents. 23 However, Gitau and colleagueshave observed impaired uterine flow in pregnant humans with documented high anxiety andattributed the impairment to possible elevations in catecholamines. 137There is some suggestion that plasma levels of catecholamines are more useful than glucocorticoidlevels for assessing stress because the former are more correlated with the intensity of thestressor. Indeed, blood catecholamine levels have been proposed as the best visceral indicator ofstress (e.g., see Natelson and coworkers 162 ). That is, catecholamine (rather than glucocorticoid)levels more accurately reflect levels of arousal. Both measures are similar in that a repeated exposureto the same neurogenic stressor results in a diminution of the response. 162 Again, these measuresrequire collection of blood, and care must be taken to avoid stressing animals during the collection.In many studies indwelling catheters are preferred for sampling, as release of catecholamines occursquickly in response to a stressor (e.g., within 5 min of the onset of restraint). Blood catecholaminelevels can be determined by high performance liquid chromatography. These methods have becomequite sophisticated in recent years and are able to detect quite small amounts (see Konarska andcoauthors 163 and Kopin 164 ).3. Stress ProteinsIn recent years there has been great interest in using those proteins known as “stress proteins” ascellular level markers for areas activated by stress or for monitoring the influence of differentenvironmental factors on animals. 165–168 The designation “stress proteins” usually includes theimmediate-early gene products (IEGPs) and the heat-shock proteins (HSPs). The HSPs were firstdiscovered in experiments involving hyperthermia but can be induced in response to other disruptive© 2006 by Taylor & Francis Group, LLC