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  • Author or Editor: Robert J. Kemppainen x
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Abstract

Objective—To determine whether inoculation of healthy dogs with a recombinant peptide containing 3 copies of ACTH would result in the production of antibodies against ACTH and whether this would affect pituitary-adrenocortical function.

Animals—8 healthy dogs.

Procedures—A recombinant peptide consisting of 3 copies of ACTH fused to a T-helper cell epitope was produced in Escherichia coli. The protein was inoculated into 4 dogs at 4-week intervals (total of 3 inoculations/dog). Four control dogs received inoculations of PBS solution mixed with adjuvant. Blood samples were collected for determination of antibody titers against ACTH and for measurement of basal and ACTH-stimulated plasma cortisol concentrations.

Results—Inoculation with the ACTH vaccine resulted in production of anti-ACTH antibodies in all 4 dogs. Titers were initially high but declined by 15 weeks after the initial inoculation. Basal cortisol concentrations were unaffected by inoculation with the ACTH vaccine. Plasma cortisol concentrations in response to ACTH stimulation were reduced at 12 weeks, but not at 15 weeks, after the first inoculation.

Conclusions and Clinical Relevance—Inoculation of dogs with a recombinant ACTH vaccine resulted in the production of antibodies against the hormone. Anti-ACTH titers were initially high but were not sustained. The only detectable endocrine effect in treated dogs was a reduction in cortisol concentration in response to ACTH stimulation in 2 of 4 dogs at 12 weeks after the first inoculation. The effect of vaccine administration on the pituitary-adrenal system was subtle and transient.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To compare results obtained from assay of total thyroxine (T4) concentration in serum of dogs and cats by use of 4 methods.

Sample Population—Serum samples obtained from 98 dogs and 100 cats and submitted by veterinarians to an endocrine testing laboratory.

Procedure—Total T4 concentration was determined in each sample by use of 4 assay methods. Assay methods included a radioimmunoassay (RIA) marketed for use in dogs, an RIA for use in humans, a chemiluminescent enzyme immunoassay for use in humans, and an in-house ELISA.

Results—Total T4 concentrations obtained by use of all methods were significantly correlated. Bias-plot comparison revealed similar good overall agreement. Total T4 concentrations determined by use of the RIA marketed for use in dogs were generally lower than concentrations measured by use of the other methods. Clinical comparisons were made by evaluation of the T4 results in the context of the reference range recommended by each laboratory. A difference was found for clinical comparisons on the basis of T4 assay method when used to identify dogs as possible hypothyroid suspects. This difference was related more to the reference range used than to the absolute T4 value. The number of hyperthyroid-suspect cats with T4 values greater than the reference range was the same for each of the 4 assay methods.

Conclusions and Clinical Relevance—Total T4 concentrations determined in dogs and cats by use of 4 commonly used methods provided similar and consistent results.

Full access
in American Journal of Veterinary Research

Abstract

Objective

To identify factors regulating secretion of α-melanocyte-stimulating hormone (α-MSH) from the pars intermedia (PI) of the pituitary gland of cats.

Animals

28 healthy adult cats.

Procedure

Indwelling catheters were placed in 1 jugular vein of each of 7 to 10 cats, depending on treatment group. Sixteen hours later, 3 blood samples were collected for determination of baseline plasma hormone concentrations, and saline solution or a test substance (haloperidol, corticotropin-releasing hormone, bromocriptine, isoproterenol, insulin, or dexamethasone) was administered via the catheter. Subsequent blood samples were collected at regular intervals for up to 240 minutes after injection. Concentrations of ACTH, cortisol, and α-MSH were measured in plasma by use of specific radioimmunoassays. Cats were rested for at least 3 weeks between experiments.

Results

Administration of haloperidol and isoproterenol resulted in increased, and bromocriptine and insulin in decreased, circulating concentrations of α-MSH from baseline. ACTH and plasma cortisol concentrations increased after administration of all test substances except dexamethasone. Dexamethasone injection resulted in decreased plasma concentrations of ACTH and cortisol.

Conclusions

Secretion of α-MSH from the PI of cats appears to be inhibited by dopaminergic activity and stimulated by β-adrenergic influences. Activation of secretion of α-MSH from the PI can be dissociated from activation of secretion of other proopiomelanocortin-derived peptides, such as ACTH, arising from the pars distalis. Regulation of secretory activity of the PI of cats resembles that of rats. (Am J Vet Res 1999;60:245–249)

Free access
in American Journal of Veterinary Research

SUMMARY

Objective

To determine whether recombinant ovine interleukin (oIL)-1 or oIL-2 alters basal or hypothalamic peptide-induced secretion of ACTH from cultured sheep pituitary cells.

Animals

The pituitary gland was collected from castrated male sheep ranging from 0.5 to 1 year old.

Procedure

Cells were cultured for 3 to 5 days, then were treated with oIL for variable periods. Cells were washed and treated with the hypothalamic peptides corticotropin-releasing hormone (CRH) or arginine vasopressin (AVP) or both. Medium bathing the cells was collected and assayed for ACTH concentration.

Results

Ovine IL-1α and oIL-1β, but not oIL-2, increased the amount of ACTH released in response to CRH, AVP, and CRH and AVP combined. Both oIL were effective after 3, but not 18 or 24 hours of exposure. Treatment with oIL-1 did not affect basal release of ACTH. Exposure of cells to phorbol 12-myristate 13-acetate or calphostin C before treatment with oIL-1β inhibited the ability of the cytokine to augment ACTH release, suggesting a role for protein kinase C in the process.

Conclusions

Local concentration of oIL-1 in the sheep pituitary gland may have an important role in determining secretion of ACTH in response to CRH or AVP or both from the hypothalamus.

Clinical Relevance

The hypothalamic-pituitary-adrenocortical axis may be activated after immune challenge. The cytokine oIL-1 has been implicated as an important mediator in this process. The pituitary gland may be an important target for this effect. (Am J Vet Res 1998;59:107–110)

Free access
in American Journal of Veterinary Research

SUMMARY

We compared the plasma Cortisol and immunoreactive corticotropin (IR-ACTH) responses to incremental doses (1.25, 12.5 and 125 μg) of synthetic ACTH (cosyntropin) administered IV to 6 clinically normal cats. Mean plasma Cortisol concentration increased significantly (P < 0-0001) after administration of all 3 doses of cosyntropin. After administration of the 1.25- and 12.5-μg doses, plasma Cortisol concentration peaked at 30 minutes, then decreased to values not significantly different from baseline concentration by 90 and 120 minutes, respectively. In contrast, after administration of the 125-μg dose, mean Cortisol concentration peaked at 60 minutes and remained significantly (P < 0.05) higher than baseline values at 120 minutes. Compared with the 1.25- and 12.5-μg doses, administration of the 125-μg dose of cosyntropin induced significantly (P < 0.05) higher Cortisol responses at 60, 90, and 120 minutes. Although individual cat's peak plasma Cortisol concentration after administration of the 125-μg dose was higher than the peak value determined after administration of the 2 lower doses of cosyntropin, these differences were not statistically significant. Mean plasma IR-ACTH concentration increased significantly (P < 0.0001) and reached a maximal value at 30 minutes after administration of all 3 doses of cosyntropin. After administration of the 1.25- and 12.5-μg doses, plasma IR-ACTH concentration decreased to values not significantly different from baseline concentration by 60 and 120 minutes, respectively, whereas mean IR-ACTH concentration remained significantly (P < 0.05) higher than baseline values 120 minutes after administration of the 125-μg dose. Mean peak plasma IR-ACTH concentration attained after administration of the 125-μg dose of cosyntropin was significantly higher than that attained after administration of the 2 lower doses. Peak plasma IR-ACTH concentration attained after administration of the 12.5-μg dose of cosyntropin was significantly higher than that attained after administration of 1.25 μg of cosyntropin. Results of the study indicate that IV administration of cosyntropin at doses ranging from 1.25 to 125 μg induces similar peak plasma cortisol responses in clinically normal cats, indicating that all of the doses may maximally stimulate the adrenal cortex. Administration of the higher cosyntropin doses did, however, result in more prolonged adrenocortical response.

Free access
in American Journal of Veterinary Research

Summary

Concentration of dexamethasone was determined in plasma or serum samples from dogs after iv administration of a low dose (0.01 mg/kg of body weight) or high dose (0.1 mg/kg) of dexamethasone. On the basis of history, clinical signs of disease, and degree of cortisol suppression in response to dexamethasone, dogs were assigned to these groups: healthy dogs, dogs with nonadrenal illness, and dogs with hyperadrenocorticism. Four hours after administration of the low dose of dexamethasone, concentration of the steroid was reduced (P < 0.05) in dogs with hyperadrenocorticism, compared with healthy dogs, but not compared with values from dogs with nonadrenal illness. By 8 hours after dexamethasone administration, values were similar across groups. Dexamethasone concentration 4 and 8 hours after high-dose administration was similar between healthy dogs and dogs with hyperadrenocorticism. Concentration of dexamethasone 4 and 8 hours after its administration overlapped after the 2 doses. For example, in 11 of 66 dogs from all groups, concentration measured 4 hours after the low dose was greater than the minimal concentration determined in the 18 dogs given the high dose. These data indicate that dexamethasone metabolism may be altered in dogs with hyperadrenocorticism, and that individuals may have appreciable variability in dexamethasone clearance. Such variability provides a possible explanation for false-positive and false-negative results associated with dexamethasone suppression testing in dogs.

Free access
in American Journal of Veterinary Research

Summary

Acid extracts of anterior and intermediate lobes of the pituitary gland from 4 dogs were fractionated by cation-exchange high-performance liquid chromatography and analzyed by radioimmunoassay, using 2 antibodies: one specific for the midregion of β-endorphin (β-end) and the other specific for the N-terminal region of N-acetylated β-end. Identification of peaks of canine β-end immunoreactivity was based on the retention times relative to those of synthetic human β-end standards, with predicted variations attributable to differences in the β-end amino acid sequences between the 2 species. The canine anterior lobe was found to contain almost exclusively β-end (1–31). By contrast, the intermediate lobe contained substantial amounts of N-acetylated and C-terminally shortened forms of β-end. The predominant forms of β-end in canine intermediate lobe were, in decreasing order of abundance: β-end (1–27), β-end (1–31), β-end (1-26), Acβ-end (1–27), Acβ-end (1–26) and Acβ-end (1–31). Individual β-end immunoreactivity profiles varied, but the general pattern was consistent among the 4 dogs.

Free access
in American Journal of Veterinary Research

SUMMARY

Plasma cortisol and immunoreactive (ir)-acth responses to 125 μg of tetracosactrin and cosyntropin—the formulation of synthetic acth available in Europe and the United States, respectively—were compared in 10 clinically normal cats. After administration of tetracosactrin or cosyntropin, mean plasma cortisol concentration reached a peak and plateaued between 60 and 120 minutes, then gradually decreased to values not significantly different from baseline concentration by 5 hours. Mean plasma ir-acth concentration reached a maximal value at 15 minutes after administration of tetracosactrin or cosyntropin and was still higher than baseline concentration at 6 hours. Difference between mean plasma cortisol and ir-acth concentrations for the tetracosactrin or cosyntropin trials was not significant at any of the sample collection times. Individual cats had some variation in the time of peak cortisol response after administration of either acth preparation. About half the cats had peak cortisol concentration at 60 to 90 minutes, whereas the remainder had the peak response at 2 to 4 hours. In general, however, peak cortisol concentration in the cats with delayed response was not much higher than the cortisol concentration at 60 to 90 minutes. Overall, these results indicate that tetracosactrin or cosyntropin induce a comparable, if not identical, pattern of adrenocortical responses when administered to healthy cats.

Free access
in American Journal of Veterinary Research

Summary

Plasma cortisol and immunoreactive (ir)-acth responses to 125 μg of synthetic acth (cosyntropin) administered iv or im were compared in 10 clinically normal cats. After im administration of cosyntropin, mean plasma cortisol concentration increased signficantly (P < 0.05) within 15 minutes, reached maximal concentration at 45 minutes, and decreased to values not significantly different from baseline concentration by 2 hours. After iv administration of cosyntropin, mean plasma cortisol concentration also increased significantly (P < 0.05) at 15 minutes, but in contrast to im administration, the maximal cortisol response took longer (75 minutes) and cortisol concentration remained significantly (P < 0.05) higher than baseline cortisol concentration for 4 hours. Mean peak cortisol concentration (298 nmol/L) after iv administration of cosyntropin was significantly (P < 0.05) higher than the peak value (248 nmol/L) after im administration. All individual peak plasma cortisol concentrations and areas under the plasma cortisol response curve were significantly (P < 0.05) higher after iv administration of cosyntropin than after im administration. Mean plasma ir-acth concentration returned to values not statistically different from baseline by 60 minutes after im administration of cosyntropin, whereas ir-acth concentration still was higher than baseline concentration 6 hours after iv administration. Peak plasma ir-acth concentration and area under the plasma ir-acth response curve also were significantly (P < 0.05) higher after iv administration of cosyntropin. Results of the study confirmed that iv adminstration of cosyntropin induces significantly (P < 0.05) greater and more prolonged adrenocortical stimulation than does im administration. The reason for this greater degree of stimulation of the adrenal cortex is presumably the result of the higher circulating acth concentration attained by iv, compared with im, administration.

Free access
in American Journal of Veterinary Research

Summary

The purpose of the study reported here was to assess 3 commonly used screening tests for hyperadrenocorticism (low-dose dexamethasone suppression test, acth stimulation test, and urinary cortisol:creatinine ratio) in dogs with various diseases other than those of the adrenal glands (nonadrenal diseases). A group of 100 dogs was studied: 59 dogs with nonadrenal disease, 21 clinically normal dogs, and 20 dogs with pituitary-dependent hyperadrenocorticism. Of 59 dogs with nonadrenal disease, 20 (34%) had high baseline cortisol concentration (greater than reference range limits), and 22 (38%) and 33 (56%) had inadequate serum cortisol suppression at 4 and 8 hours, respectively, after administration of a low dose of dexamethasone. Compared with clinically normal dogs, dogs with nonadrenal disease had significantly (P < 0.05) higher mean serum cortisol concentration at 4 and 8 hours after administration of a low dose of dexamethasone; however, significant differences were not detected between the mean cortisol concentration at 8 hours after administration for dogs with nonadrenal disease and for dogs with hyperadrenocorticism. After acth stimulation, only 8 of 59 (14%) dogs with nonadrenal disease had high serum cortisol concentrations. Significant differences did not exist after acth stimulation between mean cortisol concentration of clinically normal dogs and that of dogs with nonadrenal disease. Of 59 dogs with nonadrenal disease, 45 (76%) had a high urinary cortisol:creatinine ratio. When compared with clinically normal dogs, dogs with nonadrenal disease had a significantly higher mean urinary cortisol:creatinine ratio, but significant differences did not exist between the mean urinary cortisol:creatinine ratio of dogs with nonadrenal disease and that of dogs with hyperadrenocorticism. The calculated sensitivity for each diagnostic test used on the 20 dogs with hyperadrenocorticism in this study ranged from 0.70 to 1.0, with the acth stimulation test having the lowest sensitivity and the low-dose dexamethasone suppression test having the highest sensitivity. In dogs with nonadrenal disease, the acth stimulation test had the highest specificity (0.86) and efficiency (0.85), whereas the specificity of the low-dose dexamethasone suppression test (0.44) and urinary cortisol:creatinine ratio (0.24) were lower. The acth stimulation test also had the highest positive predictive value (0.67), whereas this value was lower for the low-dose dexamethasone suppression test (0.38) and urinary cortisol:creatinine ratio (0.25). Analysis of results of this study indicated that many dogs with nonadrenal disease have false-positive test results for hyperadrenocorticism when tested with the commonly used pituitary-adrenal function tests. Because false-positive test results were observed for all of the commonly used screening tests, a definitive diagnosis of hyperadrenocorticism should never be made solely on the basis of results of 1 or more of these screening tests, especially in dogs without classic clinical signs of hyperadrenocorticism or in dogs known to have nonadrenal disease.

Free access
in Journal of the American Veterinary Medical Association