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*Dwarfism
Endocrinology Vol. 138, No. 2 580-587
Copyright © 1997 by The Endocrine Society

Articles

Effects of Growth Hormone Secretagogues in the Transgenic Growth-Retarded (Tgr) Rat

Timothy Wells1, David M. Flavell2, Sara E. Wells, Danielle F. Carmignac and Iain C. A. F. Robinson

Division of Neurophysiology, National Institute for Medical Research, The Ridgeway, London, United Kingdom NW7 1AA

Address all correspondence and requests for reprints to: Prof. I. C. A. F. Robinson, Division of Neurophysiology, National Institute for Medical Research, The Ridgeway, Mill Hill, London, United Kingdom NW7 1AA. E-mail: i-robins{at}nimr.mrc.ac.uk


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Exogenous GH inhibits endogenous GH release by hypothalamic feedback. We have recently exploited this to generate transgenic growth-retarded (Tgr) rats, in which human GH is expressed in the hypothalamus, under the control of the rat GRF gene promoter. These rats show reduced pituitary size, GH deficiency, and dominant dwarfism, but are large enough for serial blood sampling studies to examine their spontaneous GH secretion and responses to GRF, somatostatin, and GH-releasing peptide-6 (GHRP-6). Like their normal wild-type littermates, Tgr rats show a sexually dimorphic pattern of GH secretion; males secrete GH in 3-h episodes, whereas females exhibit a more continuous irregular output, with higher baseline GH levels. In anesthetized male Tgr rats, the GH responses to GRF or GHRP-6 were markedly reduced compared with those of their nontransgenic littermates, but the differences were smaller in females. Despite the reduction in pituitary GH, peak plasma GH responses to serial GRF injections in conscious Tgr males or intermittent somatostatin infusions in conscious Tgr females were indistinguishable from the responses in their wild-type littermates. Furthermore, 7-day iv infusions of GRF (12.5–100 µg/day), given either continuously or as a pulsatile infusion stimulated growth in Tgr rats, as did pulsatile infusions of GHRP-6. Thus, despite their pituitary GH deficiency and dwarfism, Tgr rats maintain a sexually dimorphic pattern of GH release and can produce large GH secretory responses to exogenous secretagogues. They represent the first genetic model of GH deficiency in the rat in which dwarfism can be corrected by treatment with exogenous GH secretagogues.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
MAMMALIAN POSTNATAL growth is largely regulated by GH, an excess resulting in gigantism, and a deficiency in dwarfism, and some of these conditions have been modelled using various targeting strategies in transgenic mice (1, 2, 3, 4). Although overexpression of GH at peripheral sites causes enhanced growth, a dwarf phenotype results when human (h) GH is expressed in the central nervous system (5, 6). This is thought to be due to central feedback of hGH increasing hypothalamic somatostatin (SRIF) and/or reducing GRF production, which, in turn, reduces GH secretion (5, 7, 8, 9, 10, 11).

We recently generated a model of dominant GH-dependent dwarfism in the rat by expressing hGH under the control of the rat GRF promoter to direct expression specifically to hypothalamic arcuate neurons (12) and have begun to characterize the phenotype of these transgenic growth-retarded (Tgr) rats (13, 14). We chose to use rats rather than mice because dwarf rats are still large enough for their endogenous GH secretory pattern to be characterized by the chronic catheterization/microsampling technique developed for the rat (15).

Dwarf male Tgr rats continue to secrete GH in 3-h episodes, but with a reduced pulse amplitude compared to that of their wild-type littermates (12). However, despite their significant pituitary GH deficiency, growth retardation is less marked in female Tgr rats (12). The first aim of the present study was, therefore, to perform serial sampling in conscious female Tgr rats to determine whether the sexual dimorphism in the GH secretory pattern typical of normal rats (16) is maintained in this transgenic strain.

In this model of dwarfism, the pituitary should remain responsive to the normal peptidergic mechanisms controlling GH release (17). The second aim of this study was to examine the GH response to GRF, GH-releasing peptide-6 (GHRP-6) (18), or SRIF in Tgr rats, using the patterns of iv administration previously tested in normal rats (19). Due to the nature of the dwarfism, the degree of GH deficiency in the Tgr rat is less severe than that in other genetic dwarf rat strains (20, 21). The final aim of this study was, therefore, to test whether chronic infusions of GRF or GHRP-6 could stimulate sufficient endogenous GH release to correct the dwarfism in the Tgr rat. A preliminary account of some of these results has been presented (13, 14).


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Blood microsampling
All animal procedures satisfied our institutional and national ethical requirements for experiments with genetically modified rats. Hemizygous Tgr rats and their normally growing wild-type littermates from our own colony at the National Institute for Medical Research (London, UK) were generated and genotyped as previously described (12). At 7–8 weeks of age, they were placed in metabolic cages with food and water available ad libitum. After 4–10 days of acclimatization, catheters were implanted into the right jugular vein under halothane anesthesia. After at least 48 h of recovery, serial blood samples were withdrawn using a microcomputer-controlled sampling system as previously described (15). Blood samples were assayed for rat GH (rGH) by RIA either directly (15) or after separation of the plasma, and the results are expressed as nanograms per ml blood in terms of the reference preparation RP-2. Insulin-like growth factor I (IGF-I) was measured on one set of serum samples after acid-ethanol extraction (courtesy of Dr. Jenny Jones, Middlesex Hospital, London, UK).

Injection of GH secretagogues in anesthetized animals
Wild-type and Tgr male rats (6–7 weeks old; n = 6 and n = 7, respectively) were anesthetized with urethane (1.25 g/kg), and jugular vein catheters were inserted. Serial blood samples (100 µl) were taken manually before and after a bolus iv injection of 1 µg Nle27 hGRF-(1–29)NH2. After an additional 90 min, the same animals were given an iv injection of 10 µg GHRP-6 with continued manual blood sampling, and the rGH concentration was determined by RIA.

Serial GRF injections in conscious animals
Male Tgr rats and their wild-type littermates (n = 6 and n = 3, respectively) were implanted with jugular catheters under halothane anesthesia. After 48 h of recovery, animals received four consecutive injections of 1 µg [His1,Nle27]hGRF-(1–29)NH2 at 90-min intervals, during which blood samples were withdrawn automatically and assayed for rGH.

Intermittent SRIF infusions in conscious female rats
Wild-type and Tgr female rats (10–11 weeks old; n = 5 for both) were implanted with indwelling, double bore, jugular vein catheters under halothane anesthesia. After 48 h of recovery, automated blood microsampling was performed via one bore of the catheter, while a simultaneous intermittent (150 min on, 30 min off) infusion of SRIF [SRIF-(1–14); 30 µg/h] was delivered via the other bore. This procedure was previously shown to induce cycles of GH suppression and rebound secretion in conscious female rats (22). Blood samples were assayed directly for rGH.

Continuous iv infusion of GRF
Groups of 5-week old male Tgr rats (n = 3–10) were prepared with iv catheters directly connected to osmotic minipumps (Alzet model 2001, Alza Corp., Palo Alto, CA; 1 µl/h) implanted sc under halothane anesthesia. These pumps delivered PBS-BSA vehicle alone or vehicle containing [His1,Nle27]hGRF-(1–29)NH2 at a dose of 30, 60, or 100 µg/day. After 7 days the animals were weighed and killed, and the anterior pituitary was dissected, weighed, homogenized, and assayed for rGH.

Pulsatile iv infusion of GRF
Two groups of six male Tgr rats (11–12 weeks old) were equipped with indwelling jugular catheters for automatic blood sampling. After recovery, the animals were infused with 2-min (200 µl) pulses of vehicle or [His1,Nle27]hGRF-(1–29)NH2 (12.5 µg/pulse) every 3 h for 7 days (total dose, 100 µg/day) as previously described (23). After 7 days, the animals were weighed and killed just before a pulse was due. The anterior pituitary was weighed and homogenized, and the rGH content was assayed.

Pulsatile iv infusion of GHRP-6
Groups of male Tgr or wild-type rats (7–8 weeks old; n = 5 and n = 6, respectively) were equipped with indwelling jugular catheters. After recovery, a pulsatile iv infusion of GHRP-6 (10 µg in 2 min every 3 h) was continued for 7 days. Body weight and food intake were measured daily. After 7 days the animals were weighed and killed just before a pulse was due. The anterior pituitary was weighed and assayed for rGH.

Peptides used
The two GRF analogs used for these studies [Nle27]hGRF-(1–29)NH2 or [His1,Nle27]hGRF(1–29)NH2, were generous gifts from Ferring (Malmo, Sweden) and are equipotent with rGRF-(1–29)NH2 to release GH in normal rats (our unpublished data). GHRP-6 was also provided by Ferring, whereas SRIF-(1–14) was purchased from Bachem (Saffron Walden, UK). For patterned iv infusions, the peptides were dissolved in a vehicle consisting of PBS containing 0.19 mg/ml BSA and heparin (5 U/ml). For experiments with osmotic minipumps, the peptides were first dissolved at higher concentrations in 0.1 M acetic acid before dilution with the PBS/BSA vehicle, without added heparin.

Statistical analysis
Blood rGH profiles were analyzed using Pulsar (24), with the cut-off parameters set to give a false positive error rate of 5% [G(1) = 3.98; G(2) = 2.40; G(3) = 1.68; G(4) = 1.24; G(5) = 0.93]. Unless otherwise stated, pooled data are shown as the mean ± SEM, and statistical comparisons were made using Student’s t test or ANOVA followed by Bonferroni/Dunn test or Duncan’s multiple range test. A difference with P < 0.05 was considered significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
The salient characteristics of male and female Tgr rats are shown in Table 1Go. Body weight, nose-anus length, and tibial length were all significantly reduced in Tgr animals compared to those in wild-type littermates, consistent with their reduced pituitary weight, GH content, and plasma IGF-I levels. Liver, heart, and kidney weights were all significantly less in Tgr rats of both sexes (Table 1Go), but these differences largely disappeared when adjusted for their smaller total body weights. All of these effects were more pronounced in males than in females. For example, the reduction in pituitary rGH content was 75% in males compared with 43% in females, and serum IGF-I was 63% and 32% reduced in males and females, respectively, consistent with the less marked growth retardation in Tgr females.


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  		Table 1. Growth parameters, pituitary rGH, and plasma IGF-I in matched groups of 9-week-old Tgr and wild-type rats

 
Spontaneous GH secretion
Blood rGH profiles were obtained by automated microsampling from groups of five or six conscious, chronically catheterized, young adult Tgr rats and their wild-type littermates, and individual examples are shown in Fig. 1Go. The sexually dimorphic GH secretory pattern in the rat was maintained in the Tgr animals. As previously reported (12), both normal and Tgr male rats secreted GH in a regular episodic fashion, although the amounts of GH were less in the Tgr animals. In contrast, both wild-type and Tgr female rats secreted GH in a more continuous irregular GH pattern. Pulsar analysis was performed on all profiles, and the results are shown in Table 2Go. Although GH output was higher in both groups of female rats, none of the parameters of GH secretion in the Tgr females differed significantly from those in wild-type females (Table 2Go). This contrasts with the data previously obtained in male animals, which showed that GH peak height, peak areas, and total GH secretion were halved in Tgr males (analyzed in Ref. 12 and included in Table 2Go for comparison).



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  		Figure 1. Individual blood rGH profiles were obtained by automated serial sampling from conscious male and female Tgr (solid symbols) or wild-type (open symbols) rats. Analysis of these and other profiles was performed using the Pulsar program, and the results are shown in Table 2Go. Pulses identified by Pulsar are indicated in the bar above each profile.

 

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  		Table 2. Pulsar analysis of blood rGH profiles in female Tgr and wild-type rats

 
Injection of GH secretagogues in anesthetized animals
Figure 2aGo shows the effects of injecting 1 µg GRF, iv, followed by 10 µg GHRP-6, iv, in urethane-anesthetized male rats. The basal plasma rGH concentrations were significantly less in Tgr males than in their wild-type littermates. Both groups responded to GRF and GHRP-6, but the peak rGH concentrations were markedly lower in the Tgr males. Both groups also produced lower rGH responses to GHRP-6 than to GRF (note the difference in scale for GHRP-6 injections).



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  		Figure 2. The effect of GH secretagogues on plasma rGH in male (a) and female (b) urethane anesthetized wild-type (open symbols) and Tgr (closed symbols) rats. Both GRF (1 µg [Nle27]hGRF-(1–29)NH2) and GHRP-6 (10 µg) were given as bolus iv injections in 100 µl. Note the difference in scale for the GHRP-6 responses. Values shown are the mean ± SEM [n = 6 (Tgr) and n = 7 (wild-type)]. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

 
This procedure was repeated in groups of urethane-anesthetized female rats (Fig. 2bGo). In contrast to males, the peak rGH responses to GRF were similar for both Tgr and wild-type females, although the rGH response was significantly briefer in the Tgr animals. As in males, the responses to GHRP-6 were much lower than those to GRF, but the Tgr females responded at least as well as wild-type females to this peptide.

Serial GRF injections in conscious male rats
Serial iv injections of GRF were given every 90 min to groups of conscious male Tgr rats and their nontransgenic littermates (Fig. 3Go). The figure illustrates the variable rGH responses typical of male rats given GRF injections at this frequency. The first three injections showed an alternating pattern of response (the second response being significantly lower than those to the first and third injections in both groups), whereas the fourth injection elicited a GH response not significantly different from the preceding one. Notably, however, both the pattern and the amplitude of the rGH responses in these conscious male Tgr rats were indistinguishable from those in their normal littermates (Fig. 3Go).



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  		Figure 3. The effect of four serial iv injections of GRF (1 µg [His1,Nle27]hGRF-(1–29)NH2) given at 90-min intervals to conscious male Tgr (solid symbols) and wild-type rats (open symbols). Values shown are mean ± SEM [n = 6 (Tgr) or n = 3 (wild-type)]. **, P < 0.01 vs. responses 1 and 3.

 
SRIF withdrawal in conscious female rats
Alternating periods of GH suppression followed by rebound rGH secretion were seen in normal female rats given an intermittent iv infusion of SRIF (Fig. 4Go). Repeating this intermittent SRIF infusion pattern in Tgr females elicited a virtually identical pattern of rGH response. Peak rGH levels in the rebound secretory episodes were 104 ± 20 ng/ml in Tgr animal vs. 142 ± 17 ng/ml in wild-type females (P = 0.19).



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  		Figure 4. Inhibition and rebound secretion of rGH in female Tgr (solid symbols) and wild-type (open symbols) rats induced by intermittent iv infusion of SRIF (30 µg/h; 150 min on, 30 min off; bars). Values are the mean ± SEM (n = 5 for both groups).

 
Chronic iv infusions of GRF
In the first experiment, four groups of male Tgr rats were given a continuous iv infusion of vehicle or GRF at three different doses for 7 days by means of implanted osmotic minipumps. The results are shown in Fig. 5aGo. Although these 8- to 9-week-old animals were still growing relatively rapidly (>20 g/week), their growth rate could be further increased by GRF in a dose-dependent fashion. Anterior pituitary weight also increased with continuous GRF treatment, but there were no consistent or significant changes in pituitary rGH content.



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  		Figure 5. The effect of prolonged treatment of male Tgr rats with GRF. ([His1,Nle27]hGRF-(1–29)NH2) on body weight gain, anterior pituitary weight, and pituitary rGH content. GRF was administered iv either continuously from osmotic minipumps to 8- to 9-week-old rats (a) or in a pulsatile pattern (2-min pulses every 3 h) to 11- to 12-week-old rats at the doses shown for 7 days (b). Values shown are the mean ± SEM (n = 3–10/group). *, P < 0.05; ***, P < 0.001 (vs. continuously infused controls). ++. P < 0.01 (vs. pulsatile saline controls).

 
In a second experiment (Fig. 5bGo), older Tgr rats were given controlled iv infusions of saline or GRF (100 µg/day) in a pulsatile pattern (2-min pulses every 3 h for 7 days). Pulsatile GRF treatment stimulated growth markedly and increased anterior pituitary weight, again without significantly altering pituitary rGH content (Fig. 5bGo). Growth stimulation was confirmed in a further experiment with 12-week-old female Tgr rats given pulsatile GRF at a lower dose (16 µg/day; data not shown).

Pulsatile iv infusion of GHRP
As Tgr animals also responded acutely to GHRP-6 injections, the effects of chronic treatment with this secretagogue were also investigated. Figure 6Go shows an experiment in which two groups of Tgr rats were given either saline or GHRP-6 by pulsatile iv infusion (10 µg every 3 h) for 7 days and compared to an age-matched group of wild-type rats given pulsatile saline infusions. Pulsatile GHRP-6 in Tgr rats doubled their body weight gain compared with saline-infused Tgr animals, but had no significant effect on anterior pituitary weight, rGH content (Fig. 6Go), or food intake (15.5 ± 0.8 vs. 13.7 ± 0.6g/day; P = NS).



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  		Figure 6. The effect of pulsatile iv infusion of saline (200-µl pulses every 3 h) or GHRP-6 (10-µg pulses) for 7 days on body weight, anterior pituitary weight, and rGH content in male Tgr (hatched or solid bars) compared to those in a group of saline-infused wild-type rats (open bars). Values shown are the mean ± SEM [n = 5 (wild-type saline) or n = 6 (Tgr saline or GHRP-6)]. **, P < 0.01 (vs. saline-infused Tgr animals).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Following earlier observations that transgenic mice with widespread hGH expression in the central nervous system show dwarfism (5, 6), we produced a dominant dwarf transgenic rat by targeting hGH to arcuate GRF cells, which, in turn, induces somatotroph hypoplasia and reduced GH synthesis and secretion (12). The GH deficiency and dwarfism induced in this line of rats expressing a single copy dominant Tgr transgene are less severe than those in other recessive dwarf rat strains. For example, the homozygous dw/dw rat (20) has only 5–10% of the normal pituitary rGH content and grows significantly slower than the Tgr animals, whereas the homozygous SDR rat (21) cannot synthesize intact GH and is severely growth retarded.

Whole body and skeletal growth were reduced in Tgr rats of both sexes compared to those in their wild-type littermates, but the effects were more pronounced in males than in females. The 20–30% reduction in skeletal growth in hemizygote Tgr rats is accompanied by a proportionate reduction in major organ weights. Where sexual dimorphism in organ weights occurred in the wild-type littermates, it was also seen in Tgr rats for all organs except the kidney. Expression of the Tgr transgene reduced pituitary weight in both sexes, but again, the reductions in pituitary rGH content and plasma IGF-I concentrations were more pronounced in males than in females.

We have recently shown that growth retardation in Tgr males was accompanied by a significant reduction in GH secretion, although the pattern of release (12) continued to show the 3-h episodes of secretion characteristic of normal male rats (25). However, after an initial period of growth retardation, female Tgr rats grow at a rate similar to that of wild-type females (12). As the GH secretory pattern is sexually dimorphic in both normal (16) and dw/dw dwarf rats (26), we wished to test whether this was maintained in Tgr rats. The present study confirmed that female Tgr rats release GH in the continuous irregular pattern typical of normal female rats (27, 28) and showed that despite their pituitary GH deficit, the daily GH output, peak frequency, and peak height were not significantly reduced in Tgr females. This could explain their ability to achieve a normal growth rate in adulthood, unlike in males. It would be interesting to measure GH secretory profiles from females at the time their growth rate is most retarded, if the technical difficulties of serial sampling animals at 4–5 weeks (<70 g) can be overcome.

One explanation for the sex difference in dwarfism in the Tgr line might be because the major effect of the transgene is to reduce GRF output, as GRF is a major determinant of GH pulse amplitude in the male rat (29). Previous studies have also demonstrated that hGH feedback is sexually dimorphic in normal rats (30, 31). On the other hand, this sex difference was not found in a transgenic mouse line with central hGH expression (7). The difference between these two transgenic models may be due to the different pattern and extent of expression of hGH or its regulation, by the different promoters used in these models [tyrosine hydroxylase (7) vs. GRF (12)].

Although GH pulse amplitude was markedly reduced in the male Tgr rats, GH pulse frequency was not affected in either sex. This contrasts with transgenic rats with excess growth due to peripheral expression of a hGH transgene that abolished endogenous rGH pulsatility (32). Although continuous infusions of hGH also block GH pulses in normal rats (30), intermittent pulses of hGH entrain the spontaneous GH episodic rhythm in males (33), possibly by affecting the cyclic variation in hypothalamic peptide expression (34).

The GH response to GRF in Tgr rats was sex dependent and sensitive to anesthesia. The marked reduction in the peak GH responses to GRF in anesthetized Tgr males was not observed in anesthetized Tgr females, although their GH responses were significantly briefer. However, there was no reduction in the peak GH response to GRF in conscious Tgr males. As central hGH administration stimulates SRIF secretion into portal blood in urethane-anesthetized rats (11), and central hGH administration also increases SRIF expression in the hypothalamic periventricular nucleus (35), the reduced GH response to GRF seen in the anesthetized male Tgr rat could be explained if the effects of hGH to increase SRIF played a more significant feedback role in anesthetized males than in females (31). Although no differences in total hypothalamic SRIF expression were seen in their transgenic dwarf mice, Szabo et al. (7) cited other evidence to support a combined effect of hGH on both GRF and SRIF, as demonstrated previously in another transgenic mouse line (5), and we have recently found an increased level of SRIF gene expression in the periventricular nucleus in both male and female Tgr rats compared to that in their wild-type littermates (Bennett, P., I. C. A. F. Robinson, and J. Epelbaum, unpublished results).

Given their reduced pituitary GH content and spontaneous GH pulse amplitude (12), reduced GH cell number (36), and reduced GRF receptor expression (37), we expected to find a reduced GH peak response to GRF. We were, therefore, surprised to observe normal GH responses to bolus injections of GRF in conscious Tgr animals. Although GRF messenger RNA levels are reduced in Tgr rats, this was not matched by a significant reduction in GRF peptide content (12). These results can, therefore, be explained if hGH feedback reduces GRF synthesis and release, but leaves the pituitary capable of mounting a normal GH response to exogenous GRF. A discordance between spontaneous GH pulse amplitudes, and the peak GH responses to provocative testing with GRF is a well recognized problem when testing GH deficiency in human subjects (38).

Both Tgr males and their wild-type littermates showed variable responses to serial GRF injections given at 90-min intervals; the first three injections produced alternating GH responses in both wild-type and Tgr males. This confirms earlier results in normal male rats (39, 40) and has been attributed to an alternating pattern of endogenous SRIF secretion (39). If this is so, it would suggest that the pattern of endogenous SRIF is maintained in Tgr rats. To test the responsiveness to SRIF, we used a paradigm of regular intermittent infusions of SRIF, previously shown to induce cycles of GH suppression during infusion followed by large GH rebound secretory episodes after stopping the infusion (22), the amplitude of which reflects endogenous GRF release (41). Tgr rats showed normal cycles of suppression/rebound GH release when given intermittent iv infusions of SRIF, indistinguishable from the responses in their wild-type littermates. Females were chosen for this study, because the effects of intermittent SRIF are more clearly seen in them, converting their continuous endogenous GH pattern to an episodic male-type pattern (22). Intermittent SRIF infusions also induce rebound GH release in males (41), but it is more difficult to distinguish the GH rebounds from the endogenous GH secretory episodes that occur at this frequency in males (25).

Prolonged GRF treatment stimulated growth in Tgr rats and increased pituitary size, as has previously been documented in other animal models (42, 43). Other dwarf rodent strains can grow readily in response to exogenous GH or IGFs (44, 45, 46), but they do not grow in response to exogenous GH secretagogues (44, 47). For example, the dw/dw rat can release GH in response to GRF (48, 49), but in amounts insufficient to stimulate growth (47), whereas lit/lit mice are completely insensitive to acute or chronic treatment with GRF (50, 51, 52). Although both pulsatile and continuous GRF infusions were effective in Tgr rats, the magnitude of the growth responses were not directly comparable, as different doses were used in rats of different ages. Furthermore, measurements of other growth parameters (e.g. individual organ growth, skeletal growth, and IGF-I levels) will be necessary to determine the most effective dose and pattern of administration of GRF for growth promotion in these rats.

There is increasing interest in the use of novel GHRP-like secretagogues to promote growth in GH-deficient children (53), but these agents are usually tested in GH-replete models (1, 54) as most GH-deficient animal models respond poorly (20) or not at all (51) to these agents. As Tgr rats responded to GHRP-6, we also attempted to correct their dwarfism with this secretagogue. We chose pulsatile, rather than continuous, infusions of GHRP-6, because it has been shown that the growth response to a GHRP-6 analog wanes rapidly with continuous exposure (54). Tgr rats infused with pulsatile GHRP-6 grew at twice the rate of saline-infused Tgr rats and significantly faster than saline-treated nontransgenic littermates. In contrast to GRF, there was no trophic effect of GHRP-6 on the pituitary gland. This is in line with earlier studies suggesting that GHRP-6 has no direct effect on the adenylate cyclase cascade that mediates the trophic effects of GRF on the somatotroph (55, 56). Thus, the Tgr rats provides a model of GH deficiency in which the growth-promoting effects of these new classes of GH secretagogues can readily be studied.

In conclusion, sampling and infusion studies in the Tgr rat have shown that they exhibit the normal sexually dimorphic pattern of GH secretion, and that female Tgr rats release relatively normal amounts of GH despite a reduced pituitary GH reserve, which may explain their less severe dwarfism. Conscious Tgr rats respond appropriately to the hypothalamic peptides that stimulate or inhibit GH release and can produce enough GH in response to chronic treatment with GRF or GHRP-6 to correct their dwarfism.


   Acknowledgments
 
We thank Soph Sophokleous for excellent technical assistance, and Dr. Jenny Jones for IGF-I measurements. We are grateful to Ferring for the peptides used in this study, and to the NIDDK for the provision of assay reagents.


   Footnotes
 
1 Present address: Physiology Unit, MOMED, University of Wales, Cardiff, United Kingdom CF1 3US. Back

2 Present address: The Rayne Institute, University College, London, United Kingdom WC1E 6JJ. Back

Received July 29, 1996.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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