J. Anim Sci. 2007. 85:2873-2881. doi:10.2527/jas.2007-0182
© 2007 American Society of Animal Science
ANIMAL GROWTH, PHYSIOLOGY, AND REPRODUCTION |
Hormonal patterns in normal and hyperleptinemic mares in response to three common feeding-housing regimens1
W. A. Storer2,
D. L. Thompson, Jr.3,
C. A. Waller and
J. A. Cartmill
School of Animal Sciences, Louisiana Agriculture Experiment Station, Louisiana State University Agricultural Center, Baton Rouge 70803
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Abstract
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We previously reported that a rise in plasma leptin concentrations followed the rise in insulin and glucose in meal-fed horses, whereas horses maintained on pasture had little fluctuations in hormonal patterns. We have also described a hyperleptinemic-hyperinsulinemic condition that occurs in about 30% of our light horse mares of high body condition maintained on pasture. The present experiment was designed to 1) study the effect of 3 common feeding-housing regimens on leptin and other metabolic hormones in mares and 2) determine whether the hyperleptinemic condition interacted with these regimens. Six light horse mares with high body condition (average score = 7) were assigned to 2 simultaneous 3 x 3 Latin squares, 1 with normal mares (leptin = 0.1 to 6 ng/mL) and 1 with mares displaying hyperleptinemia (>10 ng/mL). Three feeding-housing regimens were compared: ad libitum pasture, ad libitum native grass hay in an outdoor paddock, and single morning feedings of a pelleted concentrate and hay at 0700 in a barn. Five days of acclimation to the feeding regimens were followed by a 36-h period of hourly blood collection to characterize the hormonal characteristics. Leptin concentrations were elevated (P < 0.001) in mares predetermined to be hyperleptinemic compared with normal mares, regardless of the feeding regimen. Leptin was greatest (P < 0.01) in mares on pasture and least in mares fed hay. Variations over time (P < 0.01) were present for all hormones and metabolites studied. Glucose and insulin concentrations were greatest (P < 0.01) in mares on pasture, with meal-fed mares exhibiting an immediate rise in plasma concentrations of both after feeding. Mares on hay had low and constant concentrations of glucose, insulin, and leptin, with no apparent fluctuations. Cortisol, prolactin, and IGF-I did not differ with leptin status, whereas GH differed due to feeding-housing regimen (P < 0.02); there was also an interaction of leptin status and feeding-housing regimen for GH concentrations (P = 0.094). It was concluded that 1) estimates of hormonal secretion in horses based on frequent sampling, depending upon the hormone in question, can be profoundly affected by the feeding-housing regimens, and 2) the hyperleptinemic condition persists under differing conditions of feeding-housing.
Key Words: feeding • hormone • housing • leptin • mare
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INTRODUCTION
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Previous studies have indicated a rise in plasma leptin concentrations approximately 8 to 10 h after a grain meal in horses (Cartmill et al., 2005
; Steelman et al., 2006). This leptin rise lagged behind the normal rises in insulin and glucose, and subsequent research (Cartmill et al., 2005) showed that insulin infusion alone, under conditions of euglycemia, caused a similar increase in leptin concentrations. In contrast to meal-fed horses, mares and geldings maintained on pasture had little change in leptin concentrations over time (Cartmill et al., 2006), unless their grazing was limited to specific periods during the day (Gentry et al., 2002a; Cartmill, 2004).
In addition to feeding effects on leptin, we have previously described a hyperleptinemic-hyperinsulinemic condition occurring in approximately 30% of our mares and geldings of high body condition maintained on pasture (Gentry et al., 2002b; Cartmill et al., 2003a). Although no effect of this condition was found in the reproductive characteristics of mares (Waller et al., 2006), possible interactions with other aspects of physiology and metabolism of the horse might be predicted based on similar syndromes in humans (Kaaja and Greer, 2005; Hotamisligil, 2007).
Often, horses in research settings are transferred back and forth from pasture to confinement in a barn to facilitate the logistics of the ongoing experiment. Although horses can be maintained long-term on hay or a combination of hay and a grain-based concentrate, the effect of these feeding regimens relative to pasture on the hormonal endpoints being measured are often unknown.
Thus, the present experiment was designed 1) to study the effect of 3 common feeding-housing regimens on leptin and other metabolic hormones in mares and 2) to determine the interaction of the hyperleptinemic condition with these regimens.
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MATERIALS AND METHODS
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All procedures involving animals in this experiment were approved by the Institutional Animal Care and Use Committee of the Louisiana State University Agricultural Center.
Mares (n = 24) in the resident herd of the Louisiana State University Agricultural Center Horse Unit were assessed during mid-March for breed, age, BCS (measured according to Henneke et al., 1983), BW, and plasma leptin concentrations. For leptin assessment, 2 samples of jugular blood were collected from each mare, 1 in the morning (approximately 0800) and 1 in the evening (approximately 1700). From the data obtained, 6 mares (Quarter Horses) were selected that had similar ages (8 to 12 yr old; mean = 11.2), BW (510 to 562 kg; mean = 542.4 kg), and BCS (6.5 to 7.5; mean = 7.0), 3 with normal leptin concentrations (0.1 to 6 ng/mL) and 3 with elevated leptin concentrations (>10 ng/mL; Cartmill et al., 2003a). The experiment was performed during late March and early April.
The experiment was performed as 2 simultaneous 3 x 3 Latin squares, 1 with the 3 normal mares and 1 with the hyperleptinemic mares. The 3 treatment (feeding-housing) regimens were as follows: 1) grass hay available ad libitum in an outdoor lot, 2) grass hay and a grain-based, pelleted concentrate fed in the morning (0700) in a stall, and 3) pasture. The pasture was predominantly winter ryegrass that had been planted the previous fall; the grass hay was predominantly Alicia Bermudagrass mixed with other native grasses on the farm that had been baled the previous summer; and the pelleted feed was Country Acres Horse Complete (Country Acres Feed Co., Brentwood, MO). Estimated compositions of the hay, concentrate, and pasture grass were 90, 93, and 30% for DM and, on a DM basis, 10, 7.8 (minimum), and 10% for CP; 2.5, 2.7, and 2.4% for crude fat; and 28, 33 (maximum), and 24% for fiber, respectively. In all feeding-housing regimens, mares were housed continuously in the feeding locations throughout that period of the Latin square (1 wk). Water was available at all times.
During each period of the Latin square, mares were acclimated to the feeding and housing regimens for 5 d (Monday through Friday). On Saturday morning, a 14-ga jugular catheter (Becton-Dickinson, Franklin Lakes, NJ) was inserted and fixed in place in each mare. After 1 h, at approximately 0700, samples of jugular blood (7 mL) were collected via the catheters every hour for 36 h (37 samples total). Blood samples were transferred to tubes containing 20 IU of sodium heparin (Sigma Chemical, St. Louis, MO) and 10 mg of NaF (Sigma) and were placed on ice until centrifuged (1,200 x g for 15 min at 5°C); plasma was harvested and stored at –20°C.
Mares in the pasture (approximately 12 ha) were free-roaming during the 5-d acclimation period except for occasional tethering on Thursday and Friday to familiarize them with the tethering process. For the blood collection period, these mares were tethered in the pasture via 10-m ropes to individual spiral anchors (tie-downs) and placed such that the mares could not become entangled with each other but could graze freely. Water was provided in buckets accessible to each mare. The anchoring positions in the pasture were moved every few hours so that ungrazed pasture was available at any given time. After dark, flashlights with red lenses were used to assist in blood sample collection. This was also the case for the other 2 treatment regimens after dark.
Mares fed hay in the outdoor lot were kept in a 20 x 25 m drylot and had ad libitum access to water (in a fixed trough) and native grass hay. They were haltered but untethered during the acclimation period as well as during the blood collection period.
Mares fed hay and a grain-based concentrate were housed individually in 4 x 4 m stalls in a barn. Each stall opened into a 4 x 12 m drylot, and mares were free to move from stall to lot during the day; they were confined to the stall after dark. Mares had ad libitum access to water in buckets hung in each stall. Each morning at approximately 0700, they were fed a pelleted commercial ration at 1% of BW (DePew et al., 1994) plus native grass hay at 1% of their BW; both the pelleted ration and the grass hay were fed on an as-fed basis. The mares were haltered but untethered during the acclimation period as well as during the blood collection period.
Concentrations of glucose in plasma were determined with a spectrophotometric, glucose oxidase-based assay (Pointe Scientific Inc., Lincoln Park, MI). Plasma concentrations of leptin (Cartmill et al., 2003a), IGF-I (Sticker et al., 1995b), GH (Thompson et al., 1992), and prolactin (Colborn et al., 1991) were assessed by RIA previously validated for horse samples. Plasma concentrations of insulin and cortisol were assessed with commercially available RIA reagents (Diagnostic Systems Laboratories, Webster, TX). Intra- and interassay CV and assay sensitivities were 6%, 4%, and 0.1 ng/mL for leptin; 5%, 12%, and 2 ng/mL for IGF-I; 8%, 11%, and 0.5 ng/mL for GH; 7%, 12%, and 0.2 ng/mL for prolactin; 5%, 8%, and 0.5 mIU/L for insulin; and 6%, 8%, and 1.1 ng/mL for cortisol.
Data for each hormone were analyzed by ANOVA as 2 simultaneous Latin squares (leptin classification, high vs. low, comprised the main squares) with repeated measures by using the GLM procedure (SAS Inst. Inc., Cary, NC). Factors in the analyses were leptin classification (normal vs. hyperleptinemic), feeding-housing regimen, horse within leptin classification, day, and sampling time. Leptin classification, feeding-housing regimen, horse within leptin classification, and day were tested with the appropriate 4-way interaction; sampling time and its interactions were tested with residual error. When mean separation was needed, the LSD test was used (Steel and Torrie, 1980).
For leptin concentrations only, additional ANOVA were performed with data from each feeding-housing regimen separately. Those ANOVA tested the effect of leptin classification, with horses within classification as the error term and time and the interaction of time and leptin classification with residual error. These ANOVA were performed to better determine whether leptin concentrations were affected by time.
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RESULTS
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Sources of variation in the ANOVA and associated P-values for each hormone and metabolite measured in the 36-h sampling period are presented in Table 1. Day was a significant source of variation for insulin (P = 0.0063), glucose (P = 0.015), and IGF-I (P = 0.02) concentrations, whereas horse within square was a significant source of variation (P < 0.02) for all but prolactin and cortisol concentrations. Because these factors were not of interest per se, and because there did not seem to be any logical trends in the daily means, they are not summarized further.
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Table 1. Sources of variation in the ANOVA and associated P-values for each hormone or metabolite measured in the 36-h sampling period1
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Leptin concentrations were affected (P < 0.01) by leptin classification and feeding-housing regimen (Table 2), with the greatest concentrations occurring when mares were grazing pasture and the least concentrations when they were consuming hay. There was an interaction (P = 0.016) of these 2 factors as well (Figure 1A), with the greatest leptin concentrations found in hyperleptinemic mares when on pasture and the lowest found in normal mares when consuming hay. There was also an effect of time (P = 0.0067) on leptin concentrations (Figure 1B), which were greatest at 6 to 8 h after the onset of sampling and again 24 h later. In the separate ANOVA performed to assess the effect of time within each feeding-housing regimen, there was a time effect (P = 0.0056) when mares were meal-fed but not (P > 0.14) when fed hay or on pasture, indicating that the pattern in Figure 1B was predominantly due to the meal-fed mares.
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Table 2. Means and SEM for the main effects of leptin classification and feeding-housing regimen for each hormone or metabolite measured in the 36-h sampling period
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Figure 1. Mean plasma concentrations of leptin in hyperleptinemic vs. normal mares for the 3 feeding-housing regimens (panel A) and averaged over all mares and feeding-housing regimens for the entire 36-h sampling period (panel B). The 3 regimens were hay (ad libitum availability of hay in an outdoor lot), meal-fed (fed a pelleted concentrate and hay, each at 1% of BW, at 0700 in a stall), and pasture (tethered in a pasture, predominantly composed of winter ryegrass). Time 0 was approximately 0700. The pooled SEM was 1.1 ng/mL for panel A and 0.9 ng/mL for panel B. a–eMeans in panel A with no like letters differ (P < 0.05).
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Insulin concentrations were affected by leptin classification (P = 0.002), feeding-housing regimen (P = 0.0021), and time (P < 0.0001). The least insulin concentrations occurred when mares consumed hay (Table 2
), and concentrations were greatest in hyperleptinemic mares. The feeding-housing regimen x time interaction (P < 0.0001; Figure 2) revealed that insulin concentrations were low and constant in mares consuming hay and high and constant in mares on pasture; when mares were meal-fed, insulin concentrations increased after each meal and remained elevated for about 6 to 8 h.
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Figure 2. Mean plasma concentrations of insulin averaged over all mares for the 3 feeding-housing regimens for the entire 36-h sampling period. The 3 regimens were hay (ad libitum availability of hay in an outdoor lot), meal-fed (fed a pelleted concentrate and hay, each at 1% of BW, at 0700 in a stall), and pasture (tethered in a pasture, predominantly composed of winter ryegrass). Time 0 was approximately 0700. The pooled SEM was 7.5 mIU/L.
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Glucose concentrations were not affected by leptin classification but were affected (P = 0.072) by feeding-housing regimen. Overall, glucose concentrations were greater in mares grazing pasture than when they consumed hay or were meal-fed (Table 2). Similar to insulin, the feeding-housing regimen x time interaction (P < 0.0001; Figure 3) revealed that glucose concentrations increased after each meal when mares were meal-fed but remained relatively constant when mares grazed pasture or consumed grass hay.
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Figure 3. Mean plasma concentrations of glucose averaged over all mares for the 3 feeding-housing regimens for the entire 36-h sampling period. The 3 regimens were hay (ad libitum availability of hay in an outdoor lot), meal-fed (fed a pelleted concentrate and hay, each at 1% of BW, at 0700 in a stall), and pasture (tethered in a pasture, predominantly composed of winter ryegrass). Time 0 was approximately 0700. The pooled SEM was 0.24 mM.
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Concentrations of IGF-I were not affected by leptin classification but were affected (P = 0.072) by feeding-housing regimen. Concentrations were greater (P < 0.1) in mares on pasture than when mares consumed hay (Table 2). There was a time effect (P = 0.0006) on IGF-I concentrations (Figure 4A), with a general trend for lower concentrations from approximately the 12th to the 20th hour of blood sampling.
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Figure 4. Mean plasma concentrations of IGF-I (panel A), prolactin (panel B), and cortisol (panel C) averaged over all mares and feeding-housing regimens for the entire 36-h sampling period. The 3 regimens were hay (ad libitum availability of hay in an outdoor lot), meal-fed (fed a pelleted concentrate and hay, each at 1% of BW, at 0700 in a stall), and pasture (tethered in a pasture, predominantly composed of winter ryegrass). Time 0 was approximately 0700. The pooled SEM were 31, 1.0, and 5.3 ng/mL for IGF-I, prolactin, and cortisol concentrations, respectively.
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Prolactin concentrations were only affected by time (P = 0.0005; Figure 4B). Examination of data of individual mares revealed considerable variation in the amount of spontaneous, pulsatile activity. However, the increases in prolactin concentrations from approximately 2 to 8 h after onset of sampling (Figure 4B), and again at 26 to 32 h, occurred during all 3 feeding-housing regimens (i.e., were observed in most mares on most occasions).
Similar to prolactin, cortisol concentrations were affected by time (P < 0.0001; Figure 4C) but no other factor. Cortisol concentrations generally declined from the onset of sampling (about 0700) and increased again beginning around 16 h into sampling (about 2300).
Concentrations of GH, which were also highly episodic in some mares on some occasions, were affected by leptin classification (P = 0.095) and feeding-housing regimen (P = 0.012); the least GH concentrations occurred when mares grazed pasture (Table 2). There was also an interaction (P = 0.094) between leptin classification and feeding-housing regimen, with GH differing (P < 0.1) between hyperleptinemic and normal mares only when fed hay (Figure 5A). The effect of time (P = 0.0002) and the interaction of feeding-housing regimen and time (P = 0.064; Figure 5B) reflected the episodic nature of GH secretion in horses.
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Figure 5. Mean plasma concentrations of GH in hyperleptinemic vs. normal mares for the 3 feeding-housing regimens (panel A) and averaged over all mares for the 3 feeding-housing regimens for the entire 36-h sampling period (panel B). The 3 regimens were hay (ad libitum availability of hay in an outdoor lot), meal-fed (fed a pelleted concentrate and hay, each at 1% of BW, at 0700 in a stall), and pasture (tethered in a pasture, predominantly composed of winter ryegrass). Time 0 was approximately 0700. a–cMeans in panel A with no like superscript differ (P < 0.1). The pooled SEM was 0.19 ng/mL for panel A and 0.94 ng/mL for panel B.
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DISCUSSION
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Although the diets that were the basis for the 3 feeding-housing regimens in the present experiment were not equivalent in nutrient content, they were typical of maintenance diets for adult horses commonly found in the horse industry. That is, they met the minimum requirements for energy, protein, Ca, P, and other nutrients, assuming they were consumed in normally expected amounts during the experiment. The experiment was not designed as a nutritional study per se but as a comparison of 3 commonly used feeding-housing regimens and their effect on hormonal and metabolite concentrations. Given that any effect of the diets would be inherently confounded with the housing condition, it is appropriate only to discuss the differences in regimens, not diets. Thus, the use of the term "when fed hay," for example, may be used for the sake of brevity but encompasses both the diet and the housing conditions, as well as any other physical and procedural factors that went along with those conditions.
Although several hormones and metabolites were measured, leptin was a primary focus of this experiment. We have been studying potential differences between mares that can be considered normal, from a leptin standpoint, and those with hyperleptinemia (Cartmill et al., 2003a), since they were first identified by Gentry et al. (2002b). In previous experiments, it was not unusual to house mares on pasture for the long-term but to confine them to stalls for periods of frequent blood sampling. As more became known about leptin in other species, particularly about short-term nutritional effects on its secretion, it became obvious that further study was necessary in horses to better define how experimental conditions may interact with other known factors, such as body fatness (Buff et al., 2002; Gentry et al., 2002b; Kearns et al., 2006) and time relative to feeding (McManus and Fitzgerald, 2000; Cartmill et al., 2005; Steelman et al., 2006).
Mares identified in the present experiment as hyperleptinemic continued to have greater leptin concentrations throughout the experiment relative to normal mares. This hyperleptinemia persisted in all 3 feeding-housing regimens, and the interaction of regimen with leptin status (Figure 1A) revealed that the hyperleptinemic state exaggerated the differences due to feeding-housing regimen. That is, for normal mares, average leptin concentrations differed only between when they were fed hay vs. when they were on pasture; for hyperleptinemic mares, all 3 regimens differed from one another. A similar exaggeration of treatment effects, and of differences between normal and hyperleptinemic mares, can be produced by short-term treatment with dexamethasone (Gentry et al., 2002a; Cartmill et al., 2003b).
The time effect on leptin concentrations was due to the variations in mares when meal-fed, because individual analyses indicated no time effect when mares were fed hay or kept on pasture. Although there was no leptin status x time interaction per se, inspection of the means for individual groups over time revealed that leptin concentrations were not just low in mares fed hay but were very constant as well. Cartmill et al. (2005) reported that meal-fed (once daily) horses had elevations in leptin concentrations 8 to 10 h after the meal. Steelman et al. (2006) reported leptin rises in horses fed twice daily (but not fed 3 or 4 times daily), and 2 other reports (Gentry et al., 2002a; Cartmill, 2004) indicated that limit-grazed horses had a similar timing of greater leptin concentrations relative to their grazing. Cartmill et al. (2005) also reported that the postmeal rise in leptin concentrations could be mimicked by insulin infusion while maintaining glucose concentrations within normal (between meal) concentrations. The fact that both insulin and leptin concentrations were relatively constant in mares fed hay and in mares on pasture in the current experiment is consistent with a model in which insulin is a major determinant of leptin secretion.
Similar to the report of Cartmill et al. (2003a), hyperinsulinemic mares had greater insulin concentrations than normal mares, and this elevation persisted across feeding-housing regimens. Although the evidence is indirect, it is likely that the continually elevated insulin concentrations are responsible for the hyperleptinemic condition of these mares. Insulin stimulates leptin secretion both in vitro (Ramsay and White, 2000; Cammisotto and Bukowiecki, 2002) and in vivo (Sivitz et al., 1998) in several species, including the horse (Cartmill et al., 2005), and leptin concentrations are correlated with fasting insulin concentrations in other species [rhesus monkey (Bodkin et al., 1996); rat (Velasque et al., 2001)]. Continually elevated insulin concentrations in humans are also associated with pathologic ovarian disease (Schroder et al., 2004), but Waller et al. (2006) reported no perturbation in winter ovarian activity or early seasonal estrous cycle characteristics in mares due to hyperleptinemia.
Insulin concentrations were greatly affected by feeding-housing regimen, with low and constant concentrations occurring throughout the 36-h period when mares were fed hay. Average concentrations were more than twice those when mares were on pasture, but again, there was little variation with time. This difference likely reflects the digestibility of the pasture grass (predominantly winter ryegrass) relative to the dry hay. Although there were the expected large increases in insulin concentrations after each meal when mares were meal-fed, between-meal concentrations were low like when mares were fed hay (which they had, depending upon when and whether they consumed their allotment from the morning).
In contrast to our earlier reports (Cartmill et al., 2003a; Waller et al., 2006), hyperleptinemic mares in the current experiment did not have elevated glucose concentrations relative to normal mares. The effect on glucose concentrations of feeding-housing regimen was primarily due to the glucose rises after each meal when mares were meal-fed. Moreover, after each rise, glucose concentrations actually dropped below the average of the other 2 treatments, such that the 36-h average was similar to that when the mares were fed hay and less than when they were on pasture. The similar glucose concentrations for mares fed hay and those on pasture, coupled with the fact that insulin concentrations were twice as high when mares were on pasture, likely indicate a high availability of soluble carbohydrates in the pasture grasses that would not be present in the hay. The moisture content of the pasture grasses could also be a major factor, because Nadal et al. (1997) reported that glucose and insulin rises after consumption of pelleted feed plus water were much greater than if water was withheld.
Interestingly, even though insulin and leptin both are elevated in hyperleptinemic mares, concentrations of IGF-I have not been found to be affected, either in the present experiment or previous experiments (Cartmill et al., 2003a). Concentrations of IGF-I are typically indicative of nutrient (particularly energy) intake of the horse in the long-term (Sticker et al., 1995b); however, Sticker et al. (1995a) reported that IGF-I concentrations did not change in mares totally deprived of feed until 32 h from the last meal. The elevation in IGF-I concentrations in mares on pasture relative to when they were fed hay likely reflects a greater energy availability or, as mentioned above, a greater availability of soluble carbohydrates in the pasture grasses that would not be present in the hay.
Prolactin concentrations were highly variable among mares due to spontaneous surges, which have been described previously (Roser et al., 1987; Thompson et al., 1994), and the increases in mean concentrations observed from the 11th to 25th hour of sampling were due solely to pastured mares. However, there was a consistent time effect across all 3 feeding-housing regimens. That is, there were rises in prolactin concentrations from 2 to 8 h after onset of blood sampling and again at 26 to 32 h in almost every individual mare, regardless of feeding-housing regimen. The prolactin response to a meal has been reported previously (DePew et al., 1994; Nadal et al., 1997), but in the absence of a meal per se, or any increase in glucose or insulin (which would indicate a period of eating after a period of fasting, if it occurred), it is not known why the mares on pasture or fed hay would have rises at this time. Other factors known to stimulate prolactin secretion in horses include exercise and stress (Thompson et al., 1988); however, there was no apparent activity or reason for anxiety observed for these mares at this time. Temperature is another possible factor that is known to alter prolactin secretion in birds (Maney et al., 1999) as well as mammals [cattle (Tucker and Wetteman, 1976); humans (Mills and Robertshaw, 1981), and sows (Barb et al., 1991)].
Cortisol concentrations were unaffected by leptin classification, which is consistent with our previous report (Cartmill et al., 2003a). They were also not affected by feeding-housing regimen, which may be a good indication that the mares were experiencing normal diurnal rhythms regardless of feeding or housing changes. Cortisol in all 3 cases varied with time, exhibiting the expected greatest concentrations early in the morning and least concentrations in the evening (Irvine and Alexander, 1994).
There were significant effects on average GH concentrations, even though the episodic nature of its secretion tends to mask changes in secretion. Similar to the report of Cartmill et al. (2003a), hyperleptinemic mares had less average GH concentrations than normal mares, albeit a small difference. Mares exhibited episodic increases in GH concentrations in all 3 feeding-housing regimens, but overall concentrations were least when mares were on pasture. Given that an increase in GH secretion has been associated with feed deprivation (Sticker et al., 1995a; Buff et al., 2005), the difference between GH concentrations in mares on pasture (less GH) and that for the other 2 feeding-housing regimens (greater GH) may reflect the difference in nutrient availability discussed above for glucose and insulin.
In summary, hyperleptinemic mares exhibited elevated insulin concentrations and slightly reduced GH concentrations relative to normal mares, and there was an interaction between leptin classification and feeding-housing regimen for each of these hormones as well. Glucose, IGF-I, prolactin, and cortisol concentrations were unaffected by leptin status, although glucose and IGF-I concentrations were affected by feeding-housing regimen. Although all 3 feeding-housing regimens are typical maintenance schemes for horses, at least in the short term, it is apparent that leptin secretion is greatly affected by the differences in the 3 regimens, as were insulin and glucose, and to a lesser degree IGF-I and GH; thus, assessments of leptin secretion in experimental protocols would need to be considered from this standpoint.
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Footnotes
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1 Approved for publication by the director of the Louisiana Agricultural Experiment Station as manuscript number 07-18-0149. We thank A. F. Parlow and the National Institute of Diabetes and Digestive and Kidney Diseases, National Hormone and Pituitary Program, Harbor-University of California Los Angeles Medical Center (Torrance, CA), and T. G. Ramsay, Growth Biology Laboratory, ARS, USDA (Beltsville, MD), for reagents. 
2 Current address: Harold and Pearl Dripps Department of Agricultural Sciences, McNeese State University, Lake Charles, LA 70609.
3 Corresponding author: dthompson{at}agctr.lsu.edu
Received for publication March 22, 2007.
Accepted for publication June 12, 2007.
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Abstract
INTRODUCTION
