J Vet Intern Med 215;29:1541 1546 Cortisol Response in Helthy nd Disesed Dogs fter Stimultion with Depot Formultion of Syntheti ACTH N.S. Sieer-Rukstuhl, W.A. Burkhrdt, N. Hofer-Inteeworn, B. Riond, I.T. Rst, R. Hofmnn-Lehmnn, C.E. Reush, nd F.S. Boretti Bkground: The ACTH stimultion test is used to evlute the drenoortil reserve. Reently, the vilility of the syntheti ACTH formultion ws limited, using mjor prolems in linil prtie. Ojetives: The ojetive of this study ws to evlute poststimultion pek ortisol onentrtions nd the durtion of the stimultory effet of depot ACTH preprtion in dogs. Animls: Twenty-two helthy dogs, 1 dogs with suspeted hypodrenoortiism (HA) nd 15 dogs with suspeted hyperdrenoortiism (HC). Methods: Prospetive study. An ACTH stimultion test using syntheti depot tetrostide, dministered intrmusulrly (5 lg/kg or t lest.1 ml) ws performed. Blood smples for determintion of ortisol were tken immeditely efore nd 1, 2, 3, 4, 6, nd 24 hours fter stimultion. Results: Pek ortisol onentrtions were rehed fter 2 4 hours in ll dogs. Cortisol onentrtions 1 hour fter stimultion were >9 lg/dl in ll helthy dogs nd >5 lg/dl in ll dogs in whih HA ws exluded. None of the dogs with HA showed ortisol-inrese ove the detetion-limit of the ssy. After 6 hours, ortisol onentrtions hd deresed in the helthy nd HC group nd were k to seline fter 24 hours. Conlusions nd Clinil Importne: The depot formultion n e used in ple of the short-ting ACTH to evlute the drenoortil reserve. Blood for pek ortisol onentrtions should e drwn 3 hours fter stimultion in ses in whih HC is suspeted; in HA-suspeted ses, lood smpling n tke ple fter 1 hour. As the stimultory effet is gone fter 24 hours, interferene with other hormonl tests is unlikely fter tht time. Key words: Adrenoortil reserve; Cnine; Synthen depot. The ACTH stimultion test hs trditionlly een reognized s n urte mesure to ssess the funtionl reserve pity of the drenl ortex nd serves s gold stndrd in dignosing hypodrenoortiism (HA) in dogs. 1,2 The test is lso used to distinguish itrogeni from spontneous hyperdrenoortiism (HC) nd to monitor trilostne or mitotne tretment. 3,4 Severl ACTH preprtions re ville. One of the most ommonly used formultions is syntheti polypeptide, whih onsists of the first 24 (of totl 39) mino ids of the ACTH, generlly known s osyntropin or tetrostide. Tetrostide is ville in 2 formultions: short-ting form for IV or IM use nd depot form for IM use only. The depot formultion ontins inorgni zin omplexes, whih dsor the From the Clini for Smll Animl Internl Mediine, (Sieer- Rukstuhl, Burkhrdt, Hofer-Inteeworn, Rst, Reush, Boretti); nd the Clinil Lortory, Vetsuisse Fulty University of Zurih, Zurih, Switzerlnd (Riond, Hofmnn-Lehmnn). The work ws done t the Clini of Smll Animl Internl Mediine, Vetsuisse Fulty University of Zurih, Zurih, Switzerlnd. Corresponding uthor: C.E. Reush, Clini for Smll Animl Internl Mediine, Vetsuisse Fulty University of Zurih, Winterthurerstrsse 26, CH-857 Zurih, Switzerlnd; e-mil: reush@vetlinis.uzh.h. Sumitted My 13, 215; Revised August 2, 215; Aepted Septemer 9, 215. Copyright 215 The Authors. Journl of Veterinry Internl Mediine pulished y Wiley Periodils, In. on ehlf of the Amerin College of Veterinry Internl Mediine. This is n open ess rtile under the terms of the Cretive Commons Attriution-NonCommeril Liense, whih permits use, distriution nd reprodution in ny medium, provided the originl work is properly ited nd is not used for ommeril purposes. DOI: 1.1111/jvim.13641 Arevitions: HA HC LDDS test PDH hypodrenoortiism hyperdrenoortiism low-dose dexmethsone suppression test pituitry-dependent hyperdrenoortiism tive sustne, resulting in protrted relese. Reently, the vilility of the short-ting tetrostide ws limited; in some ountries it ws not ville t ll. In ontrst, the vilility of the depot formultion hs never een ffeted. In humn mediine, the depot tetrostide is used for oth therpeuti nd dignosti purposes. Among other things, it hs een desried in the tretment of rheumtoid rthritis, dermtosis (ie, pemphigus, psorisis), or infntile myoloni enephlopthy. 5,6 To dignose drenl insuffiieny, the depot tetrostide is used s 5-hour test in whih ortisol onentrtions re determined efore nd every hour fter stimultion for 5-hour period. 7 Aording to the mnufturer, the durtion of the stimultory effet of the depot tetrostide n e expeted to lst etween 24 nd 36 hours. In veterinry mediine, the produt hs een evluted in helthy dogs using 2 different onentrtions (5 lg/kg nd 25 lg/dog) nd lood smpling hs een performed during the first 3 hours fter stimultion. 8 However, it is unknown whether pek serum ortisol onentrtions in dogs re rehed 3 hours fter stimultion. In ddition, the informtion s to wht time fter stimultion ortisol onentrtions re k to seline levels is lking. Knowledge out the time of pek ortisol onentrtion is prerequisite for orret
1542 Sieer-Rukstuhl et l interprettion of the test nd evlution of drenoortil reserve, nd knowing the durtion of the stimultory effet is importnt to ssess the risk of interferene with other hormonl mesurements. The use of the depot ACTH preprtion hs not yet een evluted in dogs with drenl diseses. Therefore, the ojetive of this study ws to evlute the depot tetrostide preprtion in helthy dogs nd in dogs with linil signs onsistent with HA or HC, with the ims of determining the time point of pek serum ortisol onentrtions nd the durtion of inresed ortisol onentrtions fter tetrostide depot dministrtion. Mterils nd Methods Animls Twelve purpose-red egle-pinsher mixed-reed dogs (6 femles nd 6 mles) with medin ge of 2 yers (rnge 1 2 yers) nd medin ody weight of 14.8 kg (rnge 13 17 kg) were inluded. Dogs were housed in groups in stndrd kennels t the reserh unit of the Vetsuisse Fulty of the University of Zurih, fed stndrd ommeril mintenne pellet diet one dily, nd hd d liitum ess to wter. In ddition, 1 helthy privtely owned dogs from hospitl personnel nd students, inluding 2 intt mles nd 8 femles (5 spyed), with medin ge of 5 yers (rnge 1.5 12 yers) nd medin ody weight of 15.8 kg (rnge 6.9 27 kg) were used. They were determined to e helthy on the sis of their history nd results of physil exmintion plus hemtologi nd iohemil evlution. None of the dogs hd reeived ny medition for t lest 8 weeks efore inlusion in the study exept routine vintion, deworming nd hertworm prophylxis. Ten dogs with linil suspiion of HA were prospetively enrolled. All dogs hd initilly een suspeted of hving HA sed on linil signs or lortory findings routinely seen in dogs with HA, suh s vomiting, dirrhe, wekness, lethrgy, hyperklemi nd hypontremi. Blood nd urine smples were olleted for omplete lood ount, serum iohemil profile nd urinlysis. Work-up inluded n ACTH stimultion test using the depot preprtion, whih exluded HA when post-acth ortisol onentrtion ws >5 lg/dl. Additionl tests were dominl ultrsonogrphy (in prtiulr drenl glnds) nd mesurement of endogenous ACTH. There were 5 mles (1 strted) nd 5 femles (4 spyed). Their medin ge ws 6 yers (rnge.5 14. yers) nd their medin ody weight 8.9 kg (rnge 4.9 48 kg). Fifteen dogs with linil suspiion of HC were prospetively enrolled. Dogs previously treted with gluoortioids, minerloortioids, progestgen, or trilostne were exluded from the study. They ll underwent omplete hemtologi nd iohemil evlution, urinlysis inluding urinry ulture, low-dose dexmethsone suppression test (LDDS test:.1 mg/kg dexmethsone IV, lood smpling efore, 4 nd 8 hours therefter), mesurement of endogenous ACTH nd n ultrsonogrphi exmintion of the drenl glnds. Hyperdrenoortiism ws onfirmed when the LDDS test yielded positive result (serum ortisol onentrtion 8 hours fter dexmethsone dministrtion >1. lg/dl 9 11 ) nd tretment with trilostne showed n pproprite response (strting dose of 2 mg/kg; re-evlutions performed s desried previously). Pituitry-dependent hyperdrenoortiism (PDH) ws dignosed on the sis of the dogs onentrtion of endogenous ACTH nd symmetri ultrsonogrphi pperne, with or without enlrgement, of their drenl glnds. There were 9 mles (4 strted) nd 6 femles (5 spyed). Their medin ge ws 1 yers (rnge 7 15 yers) nd their medin ody weight 13.4 kg (rnge 3.6 45. kg). Informed onsent of ll pet owners ws otined efore inluding the dogs in the study. Animl re ws in ordne with the guidelines nd diretives estlished y the Animl Welfre At of Switzerlnd. The study protool ws offiilly pproved y the veterinry offie of the nton of Zurih nd ws in ordne with the guidelines nd diretives estlished y the Animl Welfre At of Switzerlnd (TVB 191/213). Anlytil Proedures All dogs underwent n ACTH stimultion test using syntheti tetrostide hexette formultion with onentrtion of 1, lg/ml. For eh test 5 lg/kg or t lest.1 ml were dministered intrmusulrly (supr/infrspintus of the left/right thori lim or qudrieps musle of left/right hind lim) nd lood smples (jugulr venipunture) were tken immeditely efore nd 1, 2, 3, 4, 6, nd 2 24 hours fter stimultion. After lot retrtion t room temperture, serum ws hrvested y lowspeed entrifugtion nd trnsferred to tues for storge t 2 C for lter hormone ssy. Cortisol onentrtions were mesured y hemiluminesene ssy. Sensitivity of the ortisol ssy ws.2 lg/dl. All smples of eh dog were nlyzed in the sme run to minimize interssy vrition. Endogenous ACTH efore ACTH stimultion ws determined y hemiluminesene ssy. Blood ws olleted into hilled EDTA-oted tues pled on ie nd entrifuged t 4 C. Cortisol nd endogenous ACTH mesurements were performed in house twie week; plsm ws stored either t 2 C (ortisol) or t 8 C (ACTH) until ssyed. Immeditely efore nlysis, smples were thwed t room temperture. Sttistil Anlyses Dt were nlyzed with non-prmetri sttistil methods.,d Differenes etween groups of dogs were tested y using the Kruskl Wllis H-test nd the Mnn Whitney U-test. Differenes etween the time points of the ACTH stimultion were tested y use of Friedmn s repeted mesures test nd the Wiloxon mthed pirs test. All P vlues quoted re Bonferroni orreted. The level of signifine ws set t P <.5. Results No dverse retions nd no evidene of pin t the injetion site fter IM dministrtion were notied in ny of the dogs during nd fter the stimultion test. Helthy Dogs Pre- nd post-acth ortisol onentrtions of ll time points of the helthy dogs re summrized in Tle 1. There ws no signifint differene etween the vlues of helthy reserh dogs nd helthy lientowned dogs t ny time point; therefore, ll helthy dogs were summrized in 1 group for further nlysis. Mximl medin ortisol onentrtions were rehed fter 3 hours. Pek ortisol onentrtions ourred fter 1 hour in 1, fter 2 hours in 4, fter 3 hours in 11 nd fter 4 hours in 6 dogs. Twenty-four hours fter stimultion ortisol onentrtions hd gin rehed seline onentrtions. Cortisol onentrtions 1, 2, 3,
ACTH Stimultion with Depot Formultion 1543 Tle 1. Bseline nd stimulted ortisol onentrtions (medin, rnge) fter dministrtion of syntheti ACTH depot formultion. Pre-ACTH Post-ACTH hour 1 hour 2 hours 3 hours 4 hours 6 hours 24 hours Dignosis Helthy (n = 22) 1.4 (.6 2.6) 9.3 (5.1 16.4) 11.3 (5.3 24.2) 12.2,d (5.6 25.5) 9.7,,d,e (3.1 26.3) 2.7 f (.7 11.2) 1.3 (.5 3.2) Primry HA (n = 1) <.2.2 <.2 <.2 <.2 nd nd Primry HA (n = 1) <.2 <.2 <.2 <.2 <.2 nd nd Itrogeni HA (n = 1) 1.5 1.8 1.9 1.9 2.3 2.1 2.2 Non-HA (n = 7) 2 (1.5 3.7) 12.1 (9.8 18.6) 15.9,, (13.2 26.5) 17.3 (13.1 27.2) 19.8,, (2.9 3.9) 8.9,, (.7 26) 2.3,, (.4 6.4) Suspiion of HC (n = 15) 2.9 (1.9 11.5) 17. (1.6 34.1) 2.5 (7.4 37.9) 24.4,,d (3.2 34.6) 18.3,,e (1.3 29.6) 9,f (.5 2.8) 3.5 (1.8 9.6) HC (n = 1) 3.3 (1.9 11.5) 18.5 (13.8 34.1) 25.5 (15.6 37.9) 24.8,d (12.4 34.6) 24.2,,e (9.6 29.6) 16.5,f (4 2.8) 4,,,d,e (3.3 9.6) Non-HC (n = 5) 2.6 (1.9 9.9) 14.3 (1.6 21.3) 15.1 (7.4 24.6) 16.4 (3.2 24.9) 14 (1.3 14.7) 8 (.5 9) 2.7 (1.8 3.2) HC: hyperdrenoortiism; non-hc: dogs with suspiion of HC, ut negtive low-dose dexmethsone suppression test result; HA: hypodrenoortiism; non-ha: dogs with suspiion of HA, in whih HA ws exluded. Within group (row) different supersript letters indite sttistilly signifint differenes etween the time points (P <.5). 4, nd 6 hours fter stimultion were signifintly higher thn seline ortisol onentrtions (P: <.1, <.1, <.1, <.1, nd.2) nd onentrtions fter 24 hours (P: <.1, <.1, <.1, <.1, nd.2; Fig 1). Dogs with Clinil Suspiion of HA Two dogs were dignosed with primry HA (ll post- ACTH ortisol onentrtions <.2 lg/dl, ACTH > 1,25 pg/ml) nd 1 dog with itrogeni HA (ll post- ACTH ortisol onentrtions < 2.5 lg/dl, methylprednisolone pplition y privte veterinrin 4 weeks previously). In 7 dogs, HA ws exluded (post- ACTH > 5 lg/dl; non-ha group). Pre- nd post- ACTH ortisol onentrtions of the dogs with primry HA, with itrogeni HA nd with non-ha re summrized in Tle 1. In the non-ha group, mximl medin ortisol onentrtions were rehed fter 4 hours. Pek ortisol onentrtions ourred fter 2 hours in 1, fter 3 hours in 1, nd fter 4 hours in 5 dogs. Cortisol onentrtions 1 nd 3 hours fter stimultion were signifintly higher thn seline ortisol onentrtion (P =.36 nd.36; Fig 2). Dogs With Clinil Suspiion of HC Pre- nd post-acth ortisol onentrtions of the dogs with suspiion of HC re summrized in Tle 1. Mximl medin ortisol onentrtions were rehed fter 3 hours. Pek ortisol onentrtion ourred fter 1 hour in 1, fter 2 hours in 5, fter 3 hours in 6, nd fter 4 hours in 2 dogs (1 dog missing). By 24 hours fter stimultion ortisol onentrtions hd gin rehed seline onentrtion. Cortisol onentrtions 1, 2, 3, 4, nd 6 hours fter stimultion were signifintly higher thn seline ortisol onentrtions Cortisol onentrtion (ug/dl) 4 35 3 25 2 15 1 5 d 1 2 3 4 6 2-24 Hours fter ACTH dministrtion Fig 1. Serum ortisol onentrtions efore nd fter ACTH stimultion with syntheti ACTH depot formultion in helthy dogs (n = 22). Eh ox represents the interqurtile (ie, 25th to 75th perentile) rnge, the horizontl line within the ox represents the medin vlue, the rs represent the 5th to 95th perentile, nd the irles represent outlying dt points. Different letters indite sttistilly signifint differenes etween the time points (P <.5). e d f
1544 Sieer-Rukstuhl et l Cortisol onentrtion (ug/dl) 4 35 3 25 2 15 1 5 1 2 3 4 6 2-24 Hours fter ACTH dministrtion Fig 2. Serum ortisol onentrtions efore nd fter ACTH stimultion with syntheti ACTH depot formultion in dogs with initil suspiion of hypodrenoortiism (HA), ut in whih HA ws exluded (n = 7). Eh ox represents the interqurtile (ie, 25th to 75th perentile) rnge; the horizontl line within the ox represents the medin vlue; the rs represent the 5th to 95th perentile; the irles represent outlying dt points. Different letters indite sttistilly signifint differenes etween the time points (P <.5). (P =.2,.2,.2,.6, nd.26) nd onentrtions fter 24 hours (P =.1,.1,.16,.2, nd.34) (Fig 3A). In 1 dogs, HC ws onfirmed (ll PDH; HC group) nd in 5 dogs HC ws exluded (negtive LDDS test result; non-hc group). Two of these dogs were dignosed with hormonlly intive drenl mss, 1 suffered from dietes mellitus nd 2 dogs hd PU/PD of unler origin. Cortisol onentrtions t 24 hours fter stimultion were signifintly different etween the HC nd the non-hc group (P =.2; Fig 3B). Comprison etween Groups (Helthy, HC, Non-HC, Non-HA) The most distintive differenes were seen etween the helthy dogs nd the HC dogs (ll PDH). Bseline ortisol nd ortisol onentrtions 1, 2, 3, 4, 6, nd 24 hours fter stimultion were signifintly higher in dogs with HC ompred with those in helthy dogs (P: <.1, <.1, <.1, <.1,.2,.2 nd <.1; Fig 4). In the non-hc group, seline nd ortisol onentrtions 1 nd 24 hours fter stimultion (P: <.1,.2, nd.1) nd in the non-ha group, seline ortisol nd ortisol onentrtions 1 hour fter stimultion were signifintly different from those of the helthy dogs (P:.4 nd.48; Fig 4). Compring the HC nd the non-ha groups, ortisol onentrtions t 1 nd 2 hours fter stimultion were signifintly different (P =.1 nd.32; Fig 4). Between the non-hc nd the non-ha groups, no differenes were deteted. Disussion One of the gols of this study ws to evlute pek ortisol onentrtions fter stimultion with depot A Cortisol onentrtion (ug/dl) B Cortisol onentrtion (ug/dl) 4 35 3 25 2 15 1 5 4 3 2 1 1 2 3 4 6 2-24 Hours fter ACTH dministrtion HC non-hc 1 2 3 4 6 2-24 Hours fter ACTH dministrtion Fig 3. (A) Serum ortisol onentrtions efore nd fter ACTH stimultion with syntheti ACTH depot formultion in dogs with linil suspiion of hyperdrenoortiism (HC; n = 15). Eh ox represents the interqurtile (ie, 25th to 75th perentile) rnge; the horizontl line within the ox represents the medin vlue; the rs represent the 5th to 95th perentile; the irles represent outlying dt points. Different letters indite sttistilly signifint differenes etween the time points (P <.5). (B) Medin nd rnge of serum ortisol onentrtions efore nd fter ACTH stimultion with syntheti ACTH depot formultion in dogs with onfirmed HC (n = 1) nd dogs with suspiion of HC ut norml low-dose dexmethsone suppression (LDDS) test (non- HC, n = 5). *Signifint differene etween the 2 groups. tetrostide in helthy dogs nd dogs with suspiion of drenl diseses. Pek medin serum ortisol onentrtions were rehed fter 3 hours in helthy dogs nd dogs with suspeted HC nd fter 4 hours in dogs suspeted of hving HA. However, time point of pek ortisol onentrtion vried individully lying etween 1 4, 1 4, nd 2 4 hours in the helthy, non-ha group nd in dogs suspeted of hving HC, respetively. It is therefore impossile to reommend n optiml smpling time point fter stimultion with whih the pek ortisol onentrtions of ll drenl diseses n e deteted. However, onsidering the ojetive in performing n ACTH stimultion test, it is different for hypo- nd hyperfuntioning drenl glnds. In dogs suspeted of suffering from HA, the gol is either to exlude HA with d e f *
ACTH Stimultion with Depot Formultion 1545 Cortisol onentrtion (ug/dl) 4 3 2 1 # 1 2 3 4 6 2-24 Hours fter ACTH dministrtion HC non-hc non-ha Helthy iha primry HA Fig 4. Serum ortisol onentrtions efore nd fter ACTH stimultion in helthy dogs (helthy, n = 22), dogs with hyperdrenoortiism (HC, n = 1), dogs with initil suspiion of HC, ut negtive low-dose dexmethsone suppression (LDDS) test results (non-hc, n = 5), dogs with initil suspiion of hypodrenoortiism (HA), ut norml ACTH stimultion (non-ha, n = 7), dog with itrogeni HA (iha, n = 1) nd dogs with primry HA (primry HA, n = 2). For helthy, HC, non-hc nd non-ha, medin nd rnge re reported. For iha nd HA, single vlues of eh individul re reported. Sttistil omprison ws performed etween helthy, HC, non-hc nd non-ha, only. Signifint differene to helthy dogs, signifint differene to non-ha dogs. # Applies to HC, non-hc nd non-ha. ## Belongs to HC nd non-hc. suffiient stimultion (post-acth ortisol > 5 lg/dl) or to onfirm HA y n insuffiient stimultion (post- ACTH ortisol < 2 lg/dl). This gol would hve een rehed in our study, if lood hd een tken lredy 1 hour fter stimultion. All dogs suspeted of hving HA ut in whih it ws exluded hd 1-hour ortisol onentrtion of >9 lg/dl; in those with onfirmed HA it ws <.2 lg/dl. In ddition, ll helthy dogs hd 1-hour ortisol onentrtion >5 lg/dl. Therefore, it ould e onluded tht for the dignosis of HA in dogs, using the depot tetrostide, lood smple efore nd 1 hour fter stimultion with 5 lg/kg is suffiient. This informtion is of gret importne, euse if there is high suspiion of HA nd the dog is in ritil linil ondition, dministrtion of ortiosteroids should not e delyed. Exogenous syntheti ortiosteroids, however, n ross-ret with ortisol ssys or er the risk of drenl glnd suppression due to the negtive feedk mehnism, whih s onsequene would led to lunted stimultion nd misinterprettion of the test. The ojetive of n ACTH stimultion test in dogs suspeted of hving HC nd in whih trilostne tretment hs to e monitored is different. In these situtions, knowledge of the pek ortisol onentrtion fter stimultion is the importnt informtion. From our dt, lood for the dignosis of HC should e drwn etween 2 nd 4 hours fter stimultion, s within this time period the likelihood of getting the pek ortisol onentrtion is highest. Blood smpling fter 1 hour in these ses might led to flsely low test result. Bsed on our dt, no onlusion n e ## drwn s to whih time point should e hosen for monitoring the trilostne tretment, s these dogs were not evluted. It is merely speultive tht in these ses, the 1-hour vlue might lso e too erly to ssess the mximl suppression of the ortisol synthesis during trilostne tretment. Further studies re needed to ddress this issue. Another spet of pek ortisol onentrtion is the mgnitude of the pek. Two of the helthy dogs hd post-acth ortisol vlues ove the originlly estlished norml referene intervl, with vlues of 23.2 nd 26.3 lg/dl. Also, in individul dogs with suspeted HA, ortisol onentrtions of up to 3 lg/dl ould e oserved. Although these high vlues ould e interpreted s eing onsistent with HC, none of the helthy nd ertinly none of the HA-suspeted dogs hd linil signs suggestive of HC. In greement with our dt, the study of Ginel et l 8 evluting depot tetrostide in helthy Greyhounds reveled pek ortisol onentrtions with vlues up to 3 lg/dl. One might ssume tht the depot produt itself ould led to higher ortisol onentrtions ompred to the short-ting ACTH. However, study evluting short-ting ACTH for the dignosis of HA in dogs reported similr results. 12 Vlues for their dogs, using dose of 5 lg/kg, rehed ortisol onentrtions of up to 27.3 lg/dl; 1 dog even hd vlue of 41.6 lg/dl without ny evidene of HC nd with finl dignosis of inflmmtory owel disese. Therefore, we think it is more likely tht the inresed response ould hve een used y stress (white-ot effet) of the hospitliztion nd the lood smpling, or it ould e sign of the onurrent illness of the dogs. A further interesting nd importnt finding ws tht pek ortisol onentrtions rehed vlues of >22 lg/ dl in 2 of the 5 dogs where HC ws suspeted ut not onfirmed, whih ording to pulished referene vlues should e onsidered norml. 3,13,14 However, these referene vlues were estlished using older nd different ssys. Cre must e tken in dpting these referene vlues, s it is possile tht the mgnitude of pek onentrtion n reflet differenes in the ssys used to mesure ortisol. In humn mediine, it ws shown tht ortisol results from the sme ACTH test re quite different when mesured y 4 ommonly used ssys. 15 For the sme ut-off vlue, it ws possile to hve negtive test result with one nd positive result with nother ssy. The uthors onluded tht estlishing norml pek responses rely on the ssy tht is used. This is likely to e the se in veterinry mediine s well. Therefore, riteri for test interprettion proly need to e reestlished nd the ut-off level for n norml test result should e newly defined with the urrently used ssys. This hs lredy een suggested in the onsensus sttement of the ACVIM out the dignosis of spontneous nine HC. 4 A seond gol of this study ws to ssess the durtion of inresed ortisol onentrtions fter depot tetrostide dministrtion. We were le to show tht fter 24 hours the plsm ortisol onentrtions were k to seline levels in oth the helthy dogs
1546 Sieer-Rukstuhl et l nd ptient groups. This is of importne, s espeilly in dogs with suspeted HC, long durtion of inresed ortisol onentrtions fter ACTH dministrtion rries the risk of interferene with LDDS test tht my e performed the next dy. In humn mediine, plsm ACTH onentrtions hve een determined in helthy volunteers fter the dministrtion of depot ACTH. 16 While with the use of the short-ting ACTH preprtion, ACTH onentrtions dropped to undetetle levels 2 hours fter injetion, the use of the depot preprtion led to inresed plsm onentrtions up to 8 hours fter dministrtion. 16 However, levels were undetetle fter 24 hours in ll sujets. 16 This finding is somewht refleted in our dt: lthough we did not determine plsm ACTH onentrtions, we oserved sustntilly deresed ortisol onentrtion fter 6 hours s ompred to 3 hours nd, s mentioned ove, vlues were k to seline onentrtions fter 24 hours. Interestingly, dogs with onfirmed HC still hd higher ortisol onentrtions fter 24 hours ompred to dogs in whih HC ws exluded. This shows tht in dogs with hyperfuntioning drenl glnds, exogenous ACTH n result in protrted ortisol seretion in omprison with dogs with norml drenl funtion. Alterntively, it n reflet deresed ortisol lerne in dogs with HC. In onlusion, our dt show tht the depot tetrostide my e used in ple of the short-ting produt for evlution of the hypothlmi pituitry drenl xis in dogs with drenl mlfuntion. Blood smpling n tke ple 1 hour fter stimultion in dogs with suspeted HA. However, speil ttention hs to e pid in the dignosti evlution of HC, s lood smpling 1 hour fter stimultion does not revel mximl stimultion. Nonetheless, the LDDS test n e performed 24 hours fter depot tetrostide dministrtion without risk of interferene. Footnotes Synthen Depot Ò, Novrtis Phrm Shweiz AG, Bern, Switzerlnd DPC Immulite Ò 1,, Siemens Shweiz AG, Zurih, Switzerlnd GrphPd Prism6, Grph Pd Softwre, Sn Diego, CA, USA d SPSS 22. for Windows, SPSS In, Chigo, IL, USA Aknowledgments The uthors grtefully knowledge the veterinrins of the Clini for Smll Animl Internl Mediine for their ontriution of ses. We thnk ll the dog owners for their ssistne nd willingness to tke prt in this study. This study ws supported y Hill s Provet AG Switzerlnd, who kindly donted dog food for the group of helthy dogs. Conflit of Interest Delrtion: Authors dislose no onflit of interest. Off-lel Antimiroil Delrtion: Authors delre no off-lel use of ntimiroils. Referenes 1. Greo DS. Hypodrenoortiism in smll nimls. Clin Teh Smll Anim Prt 27;22:32 35. 2. Knitzer PP, Peterson ME. Primry nd seondry nine hypodrenoortiism. Vet Clin North Am Smll Anim Prt 197;27:349 357. 3. Herrtge ME, Rmsey IK. Cnine hyperdrenoortiism. In: Mooney CT, Peterson ME, eds. BSAVA Mnul of Cnine nd Feline Endorinology, 4th ed. Quedgeley, Glouester: British Smll Animl Veterinry Assoition; 212:167 189. 4. Behrend EN, Kooistr HS, Nelson R, et l. Dignosis of spontneous nine hyperdrenoortiism: 212 ACVIM onsensus sttement (smll niml). J Vet Intern Med 213;27:1292 134. 5. Ruklidge MW, Yentis SM, Peh MJ. Synthen Depot for the tretment of postdurl punture hedhe. Anesthesi 24;59:138 141. 6. Tylor WJ, Rjpkse CN, Hrris KA, et l. Inptient tretment of rheumtoid rthritis with synthen depot: A doule lind pleo ontrolled tril with 6 month followup. J Rheumtol 1999;26:2544 255. 7. Synthen Ò Depot (Tetrostidum) [pkge insert]. Rotkreuz, Shweiz: Novrtis; 212. 8. Ginel PJ, Sileo MT, Blno B, et l. Evlution of serum onentrtions of ortisol nd sex hormones of drenl glnd origin fter stimultion with two syntheti ACTH preprtions in linilly norml dogs. Am J Vet Res 212;73:237 241. 9. Burkhrdt WA, Boretti FS, Reush CE, Sieer-Rukstuhl NS. Evlution of seline ortisol, endogenous ACTH, nd ortisol/acth rtio to monitor trilostne tretment in dogs with pituitry-dependent hyperortisolism. J Vet Intern Med 213;27:919 923. 1. Qunte S, Boretti FS, Kook PH, et l. Urinry teholmine nd metnephrine to retinine rtios in dogs with hyperdrenoortiism or pheohromoytom, nd in helthy dogs. J Vet Intern Med 21;24:193 197. 11. Sieer-Rukstuhl NS, Boretti FS, Wenger M, et l. Serum onentrtions of ortisol nd ortisone in helthy dogs nd dogs with pituitry-dependent hyperdrenoortiism treted with trilostne. Vet Re 28;163:477 481. 12. Lthn P, Sott-Monrieff JC, Wills RW. Use of the ortisol-to-acth rtio for dignosis of primry hypodrenoortiism in dogs. J Vet Intern Med 214;28:1546 155. 13. Feldmn EC. Comprison of ACTH response nd dexmethsone suppression s sreening tests in nine hyperdrenoortiism. J Am Vet Med Asso 1983;182:56 51. 14. Peterson ME. Dignosis of hyperdrenoortiism in dogs. Clin Teh Smll Anim Prt 27;22:2 11. 15. Clrk PM, Neylon I, Rggtt PR, et l. Defining the norml ortisol response to the short Synthen test: Implitions for the investigtion of hypothlmi pituitry disorders. Clin Endorinol (Oxf) 1998;49:287 292. 16. Cho A, de Ceurriz J, Buisson C, et l. Simultneous quntifition nd qulifition of synthen in plsm. Anl Bionl Chem 211;399:1835 1843.