Accuracy and precision of patient positioning for pelvic MR-only radiation therapy using digitally reconstructed radiographs.

BACKGROUND AND PURPOSE: Magnetic resonance imaging (MRI) has in recent years emerged as an imaging modality to drive precise contouring of targets and organs at risk in external beam radiation therapy. Moreover, recent advances in MRI enable treatment of cancer without computed tomography (CT) simulation. A commercially available MR-only solution, MRCAT, offers a single-modality approach that provides density information for dose calculation and generation of positioning reference images. We evaluated the accuracy of patient positioning based on MRCAT digitally reconstructed radiographs (DRRs) by comparing to standard CT based workflow. MATERIALS AND METHODS: Twenty consecutive prostate cancer patients being treated with external beam radiation therapy were included in the study. DRRs were generated for each patient based on the planning CT and MRCAT. The accuracy assessment was performed by manually registering the DRR images to planar kV setup images using bony landmarks. A Bayesian linear mixed effects model was used to separate systematic and random components (inter- and intra-observer variation) in the assessment. In addition, method agreement was assessed using a Bland-Altman analysis. RESULTS: The systematic difference between MRCAT and CT based patient positioning, averaged over the study population, were found to be (mean [95% CI]) -0.49 [-0.85 to -0.13] mm, 0.11 [-0.33 to +0.57] mm and -0.05 [-0.23 to +0.36] mm in vertical, longitudinal and lateral directions, respectively. The increases in total random uncertainty were estimated to be below 0.5 mm for all directions, when using MR-only workflow instead of CT. CONCLUSIONS: The MRCAT pseudo-CT method provides clinically acceptable accuracy and precision for patient positioning for pelvic radiation therapy based on planar DRR images. Furthermore, due to the reduction of geometric uncertainty, compared to dual-modality workflow, the approach is likely to improve the total geometric accuracy of pelvic radiation therapy.

Comparison of 2 Doses for ACTH Stimulation Testing in Dogs Suspected of or Treated for Hyperadrenocorticism.

BACKGROUND: Lowering the cosyntropin dose needed for ACTH stimulation would make the test more economical. OBJECTIVES: To compare the cortisol response to 1 and 5 µg/kg cosyntropin IV in dogs being screened for hyperadrenocorticism (HAC) and in dogs receiving trilostane or mitotane for pituitary-dependent HAC. ANIMALS: Healthy dogs (n = 10); client-owned dogs suspected of having HAC (n = 39) or being treated for pituitary-dependent HAC with mitotane (n = 12) or trilostane (n = 15). PROCEDURES: In this prospective study, healthy dogs had consecutive ACTH stimulation tests to ensure 2 tests could be performed in sequence. For the first test, cosyntropin (1 µg/kg IV) was administered; the second test was initiated 4 hours after the start of the first (5 µg/kg cosyntropin IV). Dogs suspected of having HAC or being treated with mitotane were tested as the healthy dogs. Dogs receiving trilostane treatment were tested on consecutive days at the same time post pill using the low dose on day 1. RESULTS: In dogs being treated with mitotane or trilostane, the 2 doses were pharmacodynamically equivalent (90% confidence interval, 85.1-108.2%; P = 0.014). However, in dogs suspected of having HAC, the doses were not pharmacodynamically equivalent (90% confidence interval, 73.2-92.8%; P = 0.37); furthermore, in 23% of the dogs, clinical interpretation of test results was different between the doses. CONCLUSIONS AND CLINICAL RELEVANCE: For dogs suspected of having HAC, 5 µg/kg cosyntropin IV is still recommended for ACTH stimulation testing. For dogs receiving mitotane or trilostane treatment, a dose of 1 µg/kg cosyntropin IV can be used.

Characterization of the hydroxycarboxylic acid receptor 2 in cats.

The hydroxycarboxylic acid receptor 2 (HCA2) belongs to a family of nutrient-sensing receptors that bind ß-hydroxybutyrate, an alternative fuel source produced during a negative energy balance. The HCA2 receptor has not been identified or characterized in cats. Therefore, the following were the objectives of this study: (1) identify the feline HCA2 receptor protein sequence and compare against known human and rodent sequences, (2) determine tissue distribution and relative expression in lean, healthy cats, and (3) demonstrate in vitro functionality in feline adipose tissue. Tissues (n = 6) and primary adipocytes (n = 4) were collected from lean, healthy, female cats. The published genomic sequence for cats was used to design primers for polymerase chain reaction isolation of HCA2. Relative tissue distribution was evaluated using reverse transcriptase-polymerase chain reaction with RNA isolated from 9 different tissues (spleen, pancreas, lymph node, jejunum, kidney, liver, heart, and subcutaneous and abdominal adipose tissue). Receptor function was evaluated in primary feline adipocyte culture, and changes were compared with basal lipolysis. The in silico predicted feline HCA2 protein sequence exhibited 83.1% and 86.5% amino acid similarity to human and mouse sequences, respectively. The feline HCA2 receptor is predominantly expressed in adipose tissue and spleen. Exposure of feline adipocytes to niacin, a pharmacologic ligand of HCA2, inhibited lipolysis to a similar degree as insulin, a potent lipolytic inhibitor. In conclusion, the feline HCA2 receptor is similar to human and murine receptors in sequence, distribution, and functionality. By gaining a better understanding of the HCA2 receptor in cats, we will be able to better manage feline patients.

Evaluation of thyroid-stimulating hormone, total thyroxine, and free thyroxine concentrations in hyperthyroid cats receiving methimazole treatment.

BACKGROUND: Iatrogenic hypothyroidism (IH) after treatment of hyperthyroidism can impair renal function. No study compared the efficacy of measurement of serum free thyroxine by equilibrium dialysis (fT4ed) or thyroid-stimulating hormone (TSH) concentrations for monitoring cats receiving methimazole. OBJECTIVES: To (1) compare the ability of total T4 and fT4ed concentrations in conjunction with TSH to define thyroid function in hyperthyroid cats receiving methimazole, (2) determine the prevalence of IH in cats receiving methimazole, and (3) examine the relationship between thyroid axis hormones and serum creatinine concentration. ANIMALS: One hundred and twenty-five serum samples from hyperthyroid cats receiving methimazole and total T4 concentrations ≤3.9 µg/dL. METHODS: Total T4, fT4ed, and TSH concentrations were measured to evaluate thyroid status and serum creatinine concentration was measured to assess renal function. A low total T4 or fT4ed concentration in combination with an increased TSH concentration defined IH. RESULTS: Forty-one cats (33%) had increased TSH concentrations. Of cats with total T4 and fT4ed concentrations below the reference range, 68% and 73%, respectively, had TSH concentrations above the reference range. Only 18% of cats with a normal TSH concentration had an increased serum creatinine concentrations as compared to 39% of those with increased TSH concentrations (P < .001). CONCLUSIONS: Free T4ed does not identify more cats with potential IH as compared to total T4. The IH prevalence was approximately 20%. Measurement of TSH may be more helpful in indicating that azotemia, if present, is at least in part related to IH. Investigation is needed to define TSH assay utility in identifying possible subclinical IH.

Effect of trilostane and mitotane on aldosterone secretory reserve in dogs with pituitary-dependent hyperadrenocorticism.

BACKGROUND: Maximal aldosterone secretion in healthy dogs occurs 30 minutes postadrenocorticotropin (ACTH; 5 µg/kg IV) stimulation. The effect of trilostane and mitotane on aldosterone at that time is unknown. OBJECTIVES: To assess the effect of trilostane and mitotane in dogs with pituitary-dependent hyperadrenocorticism on aldosterone secretory reserve. To determine if aldosterone concentration correlates with electrolyte concentrations. ANIMALS: Serum collected from 79 client-owned dogs and 33 stored samples. METHODS: Client-owned dogs had ACTH stimulation tests with cortisol concentrations measured at 0 and 60 minutes and aldosterone concentrations measured at 0, 30, and 60 minutes. Stored samples had aldosterone concentrations measured at 0 and 60 minutes. Ten historical clinically healthy controls were included. All had basal sodium and potassium concentrations measured. RESULTS: The aldosterone concentrations in the mitotane- and trilostane-treated dogs at 30 and 60 minutes post-ACTH were significantly lower than in clinically healthy dogs; no significant difference was detected in aldosterone concentration between 30 and 60 minutes in treated dogs. However, a significantly higher percentage of dogs had decreased aldosterone secretory reserve detected at 30 minutes than at 60 minutes. At 30 minutes, decreased secretory reserve was detected in 49% and 78% of trilostane- and mitotane-treated dogs, respectively. No correlation was detected between aldosterone and serum electrolyte concentrations. CONCLUSIONS AND CLINICAL IMPORTANCE: Decreased aldosterone secretory reserve is common in trilostane- and mitotane-treated dogs; it cannot be predicted by measurement of serum electrolyte concentrations. Aldosterone concentration at 30 minutes post-ACTH stimulation identifies more dogs with decreased aldosterone secretory reserve than conventional testing at 60 minutes.

Comparison of intravenous versus intramuscular administration of corticotropin-releasing hormone in healthy cats.

BACKGROUND: Because of the lack of a current validated assay for feline endogenous adrenocorticotropic hormone (ACTH) in response to administration of currently available ovine corticotropin-releasing hormone (oCRH) preparations, a complete evaluation of the hypothalamic-pituitary-adrenal axis in cats has not been possible. OBJECTIVE: This study was undertaken to (1) determine the pituitary (ACTH) and adrenal (cortisol) response to both IV and IM administration of a currently available oCRH product in healthy cats, and (2) validate an endogenous ACTH assay for use in cats. ANIMALS: Seventeen healthy cats receiving oCRH (n = 11) or placebo (n = 6). METHODS: Prospective, randomized, placebo-controlled study. oCRH at 1 µg/kg or placebo was given either IM or IV. Endogenous cortisol and ACTH concentrations were evaluated after the injection. A comparison of IM versus IV and placebo versus treatment was made. RESULTS: The DiaSorin immunoradiometric assay (IRMA) assay for ACTH performed well, showing both parallelism and acceptable intra- and interassay coefficients of variation. There was a significant difference between groups (P = .025) and a significant difference between times (P = .025) when endogenous ACTH concentrations were compared after oCRH IV or IM. No significant differences were observed in cortisol concentrations comparing IV to IM oCRH. CONCLUSIONS: IM administration of oCRH results in significantly greater ACTH concentrations but not cortisol concentrations when compared with IV administration. Samples should be drawn before and at 60 minutes after the injection. The Diasorin IRMA is valid for feline endogenous ACTH measurements.

Seasonal changes in the combined glucose-insulin tolerance test in normal aged horses.

BACKGROUND: Equine metabolic syndrome (EMS) is an increasingly recognized problem in adult horses. Affected horses are often obese and predisposed to the development of laminitis, especially in the spring and summer months. In addition, in the summer and fall months, increases in endogenous insulin concentrations, a marker of EMS, have been reported. HYPOTHESIS/OBJECTIVES: The purpose of this study was to evaluate seasonal changes in results of the combined glucose-insulin tolerance test (CGIT), a diagnostic test for EMS. ANIMALS: Nine healthy, aged horses with no history of laminitis and no clinical signs of EMS. METHODS: Horses were given dextrose (150 mg/kg) and insulin (0.1 U/kg) IV. Plasma glucose concentrations were measured at 0, 1, 5, 15, 30, 45, 60, 75, 90, and 150 minutes and serum insulin concentrations at 0, 5, and 75 minutes. Testing was performed in February, May, June, August, September, and November. Mean glucose concentrations, characteristics of the curve, and insulin concentrations during the CGIT were compared across months using repeated measures ANOVA (P < .05). RESULTS: No CGIT parameters indicated insulin resistance, but mean area under the curve for glucose concentrations was significantly lower in August and November compared to February and in November compared to June, indicating increased insulin-mediated glucose clearance. Glucose nadir was significantly lower in November compared to that in February. CONCLUSIONS AND CLINICAL IMPORTANCE: No clinically relevant differences were seen in the results of the CGIT, suggesting that season minimally affects results of this test in normal aged horses in the southeastern United States.

Seasonal changes in plasma adrenocorticotropic hormone and α-melanocyte-stimulating hormone in response to thyrotropin-releasing hormone in normal, aged horses.

BACKGROUND: Results of diagnostic tests for equine pituitary pars intermedia dysfunction (PPID), including endogenous ACTH concentration and the overnight dexamethasone suppression test (DST), are affected by season. New and potentially more sensitive diagnostic tests for equine PPID, such as thyrotropin-releasing hormone (TRH)-stimulated ACTH response, have been developed, but have had limited evaluation of seasonality. OBJECTIVE: Our purpose was to evaluate seasonal changes in plasma ACTH and alpha-melanocyte-stimulating hormone (α-MSH) responses to TRH administration. ANIMALS: Nine, healthy, aged horses with normal DST results. METHODS: Synthetic TRH (1 mg) was administered IV. Plasma ACTH and α-MSH concentrations were measured at 0, 5, 10, 15, 20, 25, 30, 45, 60, and 180 minutes. Testing was performed in February, July, August, September, October, and November. Mean TRH-stimulated ACTH and α-MSH concentrations were compared across months and time by repeated measures analysis of variance. Significance was set at the P < .05 level. RESULTS: Concentrations of ACTH and α-MSH significantly increased after TRH administration. Endogenous and TRH-stimulated ACTH and α-MSH concentrations were significantly different across months with higher concentrations in the summer and fall compared with February. CONCLUSIONS AND CLINICAL IMPORTANCE: Plasma ACTH and α-MSH responses to TRH administration experience seasonal variation, with TRH-stimulated ACTH and α-MSH concentrations increasing from summer through fall. These results support previous evidence of a seasonal influence on the equine pituitary-adrenal axis. More research is warranted with a larger number of horses to determine if seasonal reference ranges for TRH stimulation testing need to be defined.

Pituitary-adrenal axis function in dogs with neoplasia*.

Abstract Adrenocorticotropic hormone (ACTH) stimulation tests were done in healthy and tumour-bearing dogs. In the tumour-bearing dogs, plasma endogenous ACTH (eACTH) concentration was measured and adrenal gland size was assessed ultrasonographically. Measurements in the tumour-bearing dogs were taken prior to therapy. No difference existed in basal or ACTH-stimulated cortisol concentration between tumour-bearing and healthy dogs. No difference existed in eACTH concentration between dogs with non-haematopoietic neoplasia (NHN) and lymphoma. However, of 20 dogs with lymphoma, 15% had increased basal serum cortisol concentration, 5% had an exaggerated response to ACTH and 5% had an increased eACTH concentration. Of 15 dogs with NHN, 20% had increased basal cortisol concentration, 7% had an exaggerated ACTH response and no dogs had an increased eACTH concentration. Of the dogs with lymphoma and NHN, 5 and 13%, respectively, had decreased basal cortisol concentrations; 20% of dogs with lymphoma and 13% with NHN had a subnormal ACTH response. eACTH levels were below the reference range in 10% of dogs with lymphoma and 7% with NHN. Overall, 10 adrenal glands were enlarged in seven dogs, five with lymphoma and two with NHN. The clinical significance of these findings remains to be determined.

Increased expression of MAIL, a cytokine-associated nuclear protein, in the prodromal stage of black walnut-induced laminitis.

REASONS FOR PERFORMING STUDY: The mediators and signalling cascades important in the initiation of laminitis remain unclear. We therefore wanted to explore the genes and overall signalling mechanisms that play an important role in the developmental stage of laminitis. OBJECTIVE: To use a broad genomic screening technique to identify novel genes that are differentially regulated in the equine lamellae during the developmental period of laminitis. METHODS: Differential mRNA display (DRD) was performed to discover regulated genes, and real-time quantitative polymerase chain reaction (RT-qPCR) was then used to evaluate lamellar mRNA levels of a regulated gene (MAIL) and mediators related to that gene (IL-1beta and IL-6) in control horses (n = 5) and horses administered black walnut extract (BWE; n = 5). RESULTS: Using DRD, MAIL was identified as a regulated gene. RT-qPCR indicated a 4-fold increase in expression of the MAIL mRNA in BWE lamellae compared to controls. A 30-fold increase in IL-1beta, and a 160-fold difference in IL-6 mRNA expression was present in BWE lamellae. Differences in MAIL, IL-1beta and IL-6 mRNA expression were statistically significant between groups (P < 0.05). CONCLUSIONS AND POTENTIAL RELEVANCE: The data strongly support a role for inflammatory cytokines in the developmental stages of laminitis, possibly inducing the vascular and metabolic alterations reported to occur in the affected digit. These results potentially support the use of anti-inflammatory drugs in horses at risk of laminitis, and warrant further investigation of the link between systemic disease processes associated with laminitis and the reported digital inflammation.

Regulation of Dexras1 expression by endogenous steroids.

Dexras1, a newly identified member of the Ras superfamily of proteins, was discovered in AtT-20 corticotrope cells because its expression was induced in response to glucocorticoids (dexamethasone; Dex). As yet, the function of Dexras1 is unknown, but its rapid induction in response to glucocorticoids suggests the possibility that it may be involved in negative feedback regulation of corticotropin secretion. To better understand the control of Dexras1 expression, possible effects of other steroid hormones on its expression were studied in both AtT-20 cells and in mouse pituitaries. AtT-20 cells were treated with each of 6 steroids [aldosterone, corticosterone (Cort), Dex, beta-estradiol (E(2)), progesterone and testosterone] for 2 h. Dexras1 expression was assessed using both reverse transcription polymerase chain reaction (RT-PCR) and Northern analysis. Expression of the gene was only induced in response to glucocorticoid treatment (Dex or Cort). The 6 steroids were also injected into mice, pituitaries were harvested and total RNA was obtained for RT-PCR analysis. Surprisingly, treatment with E(2), not only injection of glucocorticoids, induced Dexras1 expression in mouse pituitary. Other steroids were without effect. The results suggest that in AtT-20 corticotropes, Dexras1 expression is only induced by glucocorticoid-type steroids. In pituitary glands of mice, the gene's expression is also responsive to E(2). We conclude that either Dexras1 expression in corticotropes from normal mice is regulated differently from that in AtT-20 cells, or that Dexras1 is also expressed in other pituitary cells than corticotropes.

Diagnosis of canine hypothyroidism. Perspectives from a testing laboratory.

The most common sample received by our endocrine testing laboratory is submitted for the diagnosis of hypothyroidism in a dog. The current tests most frequently employed in our laboratory for thyroid evaluation in dogs are total T4, free T4 by dialysis, and canine TSH measurement. Each test has strengths and weaknesses and suffers from the possibility of both false positive and false negative results. This article provides a working description of each test and an approach to interpretation of results. Other tests that are less commonly used are also discussed. Examples of interpretation of test results in individual hypothyroid-suspect dogs are presented for illustration.

Diagnosis of canine hyperadrenocorticism.

Canine hyperadrenocorticism is one of the most common endocrinopathies in dogs. Diagnosis remains difficult in some cases due to factors such as the presence of non-adrenal illness and limitations in the tests. Differentiation between the pituitary and adrenal forms is important for providing accurate prognostic information and delineating treatment options and protocols. This article reviews the tests available for diagnosis (screening) and differentiation and evaluates their advantages and disadvantages. Recommendations for testing are made.

Reduced growth of calves and its reversal by use of anabolic agents.

Disease has profound effects on the immune system, endocrine system, and on the growth process. Since diseases are catabolic to the animal, there is current interest in the possible role of anabolic hormones to counter the effects of disease in general and minimize the effects of a disease process on growth and development. A number of anabolic hormones, such as growth hormone (GH) and estradiol + progesterone (EP), have been studied for their role in enhancing growth and stimulating immune function and are thus candidates for hormonal intervention in disease processes. GH has been shown to be effective in countering some of the deleterious effects of endotoxemia but was ineffective in a parasitic disease model. Studies with EP have shown similar success with both endotoxemia and a parasitic disease model. Moreover, GH and EP do not share a common mechanism of action, suggesting that the effects are not simply due to anabolic actions. While the mechanism of action of GH in endotoxemia has been examined, the effects of EP are via an unknown mechanism, possibly by inhibition of IL-I action or inhibition of nitric oxide overproduction.

Effect of alternate-day prednisolone administration on hypophyseal-adrenocortical activity in dogs.

OBJECTIVE: To evaluate effect of alternate-day oral administration of prednisolone on endogenous plasma ACTH concentration and adrenocortical response to exogenous ACTH in dogs. ANIMALS: 12 Beagles. PROCEDURE: Dogs were allotted to 2 groups (group 1, 8 dogs treated with 1 mg of prednisolone/kg of body weight; group 2, 4 dogs given excipient only). During a 30-day period, blood samples were collected for determination of plasma ACTH and cortisol concentrations before, during, and after treatment with prednisolone. From day 7 to 23, prednisolone or excipient was given on alternate days. Sample collection (48-hour period with 6-hour intervals) was performed on days 1, 7, 15, 21, and 28; on other days, sample collection was performed at 24-hour intervals. Pre- and post-ACTH plasma cortisol concentrations were determined on days 3, 9, 17, 23, and 30. RESULTS: A significant difference was detected between treatment and time for group 1. Plasma ACTH concentrations significantly decreased for 18 to 24 hours after prednisolone treatment in group-1 dogs. At 24 to 48 hours, ACTH concentrations were numerically higher but not significantly different in group-1 dogs. Post-ACTH plasma cortisol concentration significantly decreased after 1 dose of prednisolone and became more profound during the treatment period. However, post-ACTH cortisol concentration returned to the reference range 1 week after prednisolone administration was discontinued. CONCLUSIONS AND CLINICAL RELEVANCE: Single oral administration of 1 mg of prednisolone/kg significantly suppressed plasma ACTH concentration in dogs for 18 to 24 hours after treatment. Alternate-day treatment did not prevent suppression, as documented by the response to ACTH.

Evaluation of a low-dose synthetic adrenocorticotropic hormone stimulation test in clinically normal dogs and dogs with naturally developing hyperadrenocorticism.

OBJECTIVE: To determine whether low doses of synthetic ACTH could induce a maximal cortisol response in clinically normal dogs and to compare a low-dose ACTH stimulation protocol to a standard high-dose ACTH stimulation protocol in dogs with hyperadrenocorticism. DESIGN: Cohort study. ANIMALS: 6 clinically normal dogs and 7 dogs with hyperadrenocorticism. PROCEDURE: Each clinically normal dog was given 1 of 3 doses of cosyntropin (1, 5, or 10 micrograms/kg [0.45, 2.3, or 4.5 micrograms/lb] of body weight, i.v.) in random order at 2-week intervals. Samples for determination of plasma cortisol and ACTH concentrations were obtained before and 30, 60, 90, and 120 minutes after ACTH administration. Each dog with hyperadrenocorticism was given 2 doses of cosyntropin (5 micrograms/kg or 250 micrograms/dog) in random order at 2-week intervals. In these dogs, samples for determination of plasma cortisol concentrations were obtained before and 60 minutes after ACTH administration. RESULTS: In the clinically normal dogs, peak cortisol concentration and area under the plasma cortisol response curve did not differ significantly among the 3 doses. However, mean plasma cortisol concentration in dogs given 1 microgram/kg peaked at 60 minutes, whereas dogs given doses of 5 or 10 micrograms/kg had peak cortisol values at 90 minutes. In dogs with hyperadrenocorticism, significant differences were not detected between cortisol concentrations after administration of the low or high dose of cosyntropin. CLINICAL IMPLICATIONS: Administration of cosyntropin at a rate of 5 micrograms/kg resulted in maximal stimulation of the adrenal cortex in clinically normal dogs and dogs with hyperadrenocorticism.

Regulation of alpha-melanocyte-stimulating hormone secretion from the pars intermedia of domestic cats.

OBJECTIVE: To identify factors regulating secretion of alpha-melanocyte-stimulating hormone (alpha-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 alpha-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 alpha-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 alpha-MSH from the PI of cats appears to be inhibited by dopaminergic activity and stimulated by beta-adrenergic influences. Activation of secretion of alpha-MSH from the PI can be dissociated from activation of secretion of other pro-opiomelanocortin-derived peptides, such as ACTH, arising from the pars distalis. Regulation of secretory activity of the PI of cats resembles that of rats.

Effect of storage conditions on cortisol, total thyroxine, and free thyroxine concentrations in serum and plasma of dogs.

OBJECTIVE: To determine for dogs stability of cortisol, thyroxine (T4), and free thyroxine (fT4) in plasma and serum stored in glass or plastic tubes at -20, 4, 25, and 37 C. DESIGN: Prospective study. ANIMALS: Phase I, 7 Greyhounds; Phase II, 6 mixed-breed dogs. PROCEDURE: Phase I: blood was obtained after administration of thyroid-stimulating hormone and adrenocorticotropin. Serum and plasma samples from each dog were divided into 8 aliquots, 4 in glass and 4 in plastic tubes. A pair of aliquots, 1 in plastic and 1 in glass, were stored at -20, 4, 25, or 37 C for 5 days and then assayed for hormones. Phase II: blood was obtained without prior stimulation. For fT4 determination, serum from each dog was placed in plastic or glass tubes, assayed immediately, stored at -20 C for 5 days, and reassayed. Aliquots from each dog were also stored for 1 day at 4 or 25 C and then assayed. Samples for cortisol determination were handled as in phase I. RESULTS: Phase I: there was no effect of tube type (glass vs plastic) on cortisol. Cortisol concentrations decreased after storage in serum at 4, 25, and 37 C, and in plasma at 37 C, compared with storage at -20 C. There was no effect of sample type (serum or plasma) on T4. Thyroxine concentrations increased after storage at 37 C in glass, compared with storage at -20 C. The fT4 concentrations were lower in serum than plasma after storage at -20 C. Concentrations of fT4 increased after storage at 37 C in glass, compared with storage at -20 C. Phase II: the fT4 concentrations did not change after storage in any condition. There was no effect of tube type on cortisol concentrations. Serum cortisol concentrations decreased after storage at 37 C, compared with storage at -20 C. CLINICAL IMPLICATIONS: For cortisol, cooling of plasma is not necessary, but serum should be shipped cold. For T4 and fT4, serum is sufficient; contained within plastic tubes, samples can be shipped without cooling if assayed within 5 days.

Dexamethasone rapidly induces a novel ras superfamily member-related gene in AtT-20 cells.

Differential display was used to identify a new Ras superfamily gene (Dexras1) induced by dexamethasone (Dex) in AtT-20 cells. Treatment of AtT-20 cells with Dex for 30 min resulted in increased mRNA for Dexras1; the highest concentrations appeared after 2 h of treatment. The gene was also identified in mouse heart, brain, liver, and kidney and furthermore was induced in these tissues after Dex treatment. The deduced protein shows regions of homology characteristic of members of the Ras superfamily of small GTPases. Highest homology (36% identity, 57% positives) was found with human Rap-2b, followed closely by a number of other Ras subfamily members, suggesting that Dexras1 is probably a member of the Ras subfamily of GTPases (members include Ras and Rap). Dexras1 is the first Ras superfamily member identified that is induced in response to steroids. The function of this gene is unknown; however, its wide distribution and rapid induction by Dex suggests the possibility of a role in glucocorticoid action in a variety of tissues.