Ethyl Amide Research Articles

Bimatoprost, a novel efficacious ocular hypotensive drug now recognized as a member of a new class of agents called prostamides

AbstractPursuit of a new FP‐agonist prodrug led to the identification of an interesting series of neutral C1‐substituted prostaglandin F2α analogues. Although these initial analogues were devoid of any inherent pharmacological activity at the FP‐receptor, two compounds AGN‐190910 and AGN‐191129, were found to have pronounced effects in lowering intraocular pressure (IOP) in normotensive dogs and monkeys. The cat iris sphincter assay was quickly developed as a primary screen for these analogues, leading to rapid identification of AGN‐192024 (17‐phenyl PGF2α ethyl amide, bimatoprost). While bimatoprost is structurally similar to naturally occurring mammalian hormones of the prostanoid family, surprisingly it demonstrates no significant activity at any of the known prostanoid receptors. Furthermore, results of considerable additional pharmacological studies provide evidence that it may indeed act through a unique receptor yet to be identified. The effect of Bimatoprost on lowering IOP has also been found to be unique in comparison to prostanoids. Bimatoprost reduces human IOP by increasing aqueous humor outflow through a dual mechanism of action where it improves both pressure‐dependent and pressure‐independent outflow pathways. First introduced to the market in 2002, bimatoprost is currently the most potent single therapy available for control of ocular hypertension. Drug Dev Res 68:147–155, 2007. ©2007 Wiley‐Liss, Inc.

The Synthetic Triterpenoids CDDO-Methyl Ester and CDDO-Ethyl Amide Prevent Lung Cancer Induced by Vinyl Carbamate in A/J Mice

We report the first use of new synthetic triterpenoids to prevent lung cancer in experimental animals. Female A/J mice were treated with the mutagenic carcinogen vinyl carbamate, which induces adenocarcinoma of the lung in all animals within 16 weeks. If mice were fed either the methyl ester or the ethyl amide derivative of the synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO-ME and CDDO-EA, respectively), beginning 1 week after dosing with carcinogen, the number, size, and severity of lung carcinomas were markedly reduced. The mechanisms of action of CDDO-ME and CDDO-EA that are germane to these in vivo findings are the following results shown here in cell culture: (a) suppression of the ability of IFN-gamma to induce de novo formation of nitric oxide synthase in a macrophage-like cell line RAW264.7, (b) induction of heme oxygenase-1 in these RAW cells, and (c) suppression of phosphorylation of the transcription factor signal transducers and activators of transcription 3 as well as induction of apoptosis in human lung cancer cell lines.

4-Hydroxy-2-quinolones. 107. Reaction of triethyl methanetricarboxylate with indoline

The first stage of the reaction of triethyl methanetricarboxylate with indoline is the formation of the diethyl ester of 2-(indoline-1-carbonyl)malonic acid, which then, depending on the conditions selected, may be converted into the ethyl ester of 2-(indoline-1-carbonyl)-3-(indolin-1-yl)-3-oxopropionic acid, methanetri-N-(indolin-1-yl)carboxamide, or the ethyl ester or (indolin-1-yl)amide of 1-hydroxy-3-oxo-5,6-dihydro-3H-pyrrolo[3,2,1-ij]quinoline-2-carboxylic acid.

Thyroid receptor ligands. Part 2: Thyromimetics with improved selectivity for the thyroid hormone receptor beta

A set of thyromimetics having improved selectivity for TR-β1 were prepared by replacing the 3 ′-isopropyl group of 2 and 3 with substituents having increased steric bulk. From this limited SAR study, the most potent and selective compounds identified were derived from 2 and contained a 3 ′-phenyl moiety bearing small hydrophobic groups meta to the biphenyl link. X-ray crystal data of 15c complexed with TR-β1 LBD shows methionine 442 to be displaced by the bulky R3 ′ phenyl ethyl amide side chain. Movement of this amino acid side chain provides an expanded pocket for the bulky side chain while the ligand–receptor complex retains full agonist activity.

Synthesis of prostaglandin F ethanolamide by prostaglandin F synthase and identification of Bimatoprost as a potent inhibitor of the enzyme: new enzyme assay method using LC/ESI/MS

Prostaglandin (PG) D 2 ethanolamide (prostamide D 2) was reduced to 9α,11β-PGF 2 ethanolamide (9α,11β-prostamide F 2) by PGF synthase, which also catalyzes the reduction of PGH 2 and PGD 2 to PGF 2α and 9α,11β-PGF 2, respectively. These enzyme activities were measured by a new method, the liquid chromatographic–electrospray ionization–mass spectrometry (LC/ESI/MS) technique, which could simultaneously detect the substrate and all products. PGF 2α, 9α,11β-PGF 2, PGD 2, PGH 2, 9α,11β-prostamide F 2, and prostamide D 2 were separated on a TSKgel ODS 80Ts column, ionized by electrospray, and detected in the negative mode. Selected ion monitoring (SIM) of m/ z 353 ([M–H] −), 353 ([M–H] −), 351 ([M–H] −), 333 ([M–H–H 2O] −), 456 ([M+59] −), and m/ z 358 ([M−37] −) was used for quantifying PGF 2α, 9α,11β-PGF 2, PGD 2, PGH 2, 9α,11β-prostamide F 2, and prostamide D 2, respectively. The detection limit for PGF 2α and 9α,11β-PGF 2 was 0.01 pmol; that for PGH 2 and PGD 2, 0.1 pmol; and that for prostamide D 2 and 9α,11β-prostamide F 2, 0.5 and 0.03 pmol, respectively. The LC/ESI/MS technique for measuring PGF synthase activity showed higher sensitivity than other methods. Using this method, we found that Bimatoprost, the ethyl amide analog of 17-phenyl-trinor PGF 2α and an anti-glaucoma agent, inhibited all three reductase activities of PGF synthase when used at a low concentration. These results suggest that Bimatoprost also behaves as a potent PGF synthase inhibitor in addition to having prostamide-like activity.

The Hydrolysis of the Prostaglandin Analog Prodrug Bimatoprost to 17-Phenyl-trinor PGF2α by Human and Rabbit Ocular Tissue

Bimatoprost (Lumigan), the ethyl amide derivative of the potent prostaglandin FP agonist 17-phenyl-trinor PGF(2alpha), has been reported to be a member of a pharmacologically unique class of ocular hypotensive agents. To confirm that bimatoprost, which is intrinsically active as an FP prostaglandin agonist, is also a prostaglandin analog prodrug, the hydrolysis of bimatoprost by ocular tissues was studied by incubating solutions containing bimatoprost with either human or rabbit ocular tissue. The ethyl amide group of bimatoprost was hydrolyzed by rabbit and human cornea, iris/ciliary body and Thasclera to produce the expected carboxylic acid product, 17-phenyl-trinor PGF(2alpha). The rate of hydrolysis by human and rabbit cornea and iris/ciliary body is similar, whereas the rate of hydrolysis by the sclera is slower in humans than in rabbits. These studies show that human and rabbit ocular tissue (cornea, iris/ciliary body and sclera) can convert bimatoprost to the potent prostaglandin FP agonist 17-phenyl-trinor PGF(2alpha). Separate in vitro studies clearly show that both bimatoprost and 17-phenyl-trinor PGF(2alpha) have affinity for and are agonists at the human FP receptor. Taken together, the data strongly suggests that the ocular hypotensive effect of bimatoprost can be attributed to its activity as a prostaglandin receptor agonist either directly or through its role as a prostaglandin agonist prodrug.

Real-time intracellular Ca2+ mobilization by travoprost acid, bimatoprost, unoprostone, and other analogs via endogenous mouse, rat, and cloned human FP prostaglandin receptors.

The ability of a number of prostaglandin F 2 alpha (PGF 2 alpha) analogs to mobilize intracellular Ca2+[Ca2+]iand to compete for [3H]PGF 2 alpha binding to prostaglandin F 2 alpha receptors (FP) was evaluated. Radioligand binding studies measuring displacement of [3H]PGF 2 alpha by a variety of FP prostaglandin analogs yielded the following rank order of affinities: travoprost acid [(+)-16-m-trifluorophenoxy tetranor PGF 2 alpha; (+)-fluprostenol] > bimatoprost acid (17-phenyl-trinor PGF 2 alpha) >> unoprostone (13,14-dihydro-15-keto-20-ethyl PGF 2 alpha) = bimatoprost (17-phenyl-trinor PGF 2 alpha ethyl amide) > or = Lumigan (bimatoprost ophthalmic solution). In FP functional studies, travoprost acid (EC50= 17.5-37 nM, n = 13), bimatoprost acid (EC50= 23.3-49.0 nM, n = 6-12), unoprostone (EC50= 306-1270 nM, n = 4-8), bimatoprost (EC50= 3070- 3940 nM, n = 4-9), and Lumigan (EC50= 1470-3190 nM, n = 5-9) concentration dependently stimulated [Ca2+]imobilization via the rat (A7r5 cells), mouse (3T3 cells), and cloned human ocular FP prostanoid receptors. The rank order of potency of these compounds at the FP receptor of the three species was similar and in good agreement with the determined binding affinities. The agonist effects of these compounds were concentration dependently blocked by the FP receptor-selective antagonist, AL-8810 (11beta-fluoro-15-epi-15-indanyl-tetranor PGF 2 alpha) (Ki= 0.6-1.3 microM). These studies have demonstrated that bimatoprost, unoprostone, and bimatoprost acid possess direct agonist activities at the rat, mouse, and human FP prostanoid receptor and that travoprost acid is the most potent of the synthetic FP prostaglandin analogs tested.

Bimatoprost (Lumigan ®) is an agonist at the cloned human ocular FP prostaglandin receptor: real-time FLIPR-based intracellular Ca 2+ mobilization studies

Bimatoprost is the ethyl amide derivative of 17-phenyl-trinor prostaglandin F 2 α . Here, we show that bimatoprost ( K i =9250±846 nM) and bimatoprost free acid (17-phenyl-trinor prostaglandin F 2 α ; K i =59±6 nM) bind to the FP receptor and displace [ 3H]-travoprost acid, a selective FP agonist. Bimatoprost (EC 50=3070±1330 nM), Lumigan ® (bimatoprost 0.03% ophthalmic solution; EC 50=1150±93 nM) and bimatoprost acid (EC 50=15±3 nM) mobilized intracellular Ca 2+ ([Ca 2+] i ) in <5 s in HEK-293 cells expressing the cloned human ciliary body FP receptor on a fluorometric imaging plate reader (FLIPR). Furthermore, agonist effects of bimatoprost and bimatoprost acid were blocked by AL-8810 (11 β-fluoro-15-epi-15-indanyl prostaglandin F 2 α ; K i =0.7–2.1 μM), an FP receptor-selective antagonist. Therefore, the prodrug bimatoprost and its hydrolytic product, bimatoprost free acid, bind to and activate the human ocular FP prostaglandin receptor to mobilize [Ca 2+] i , thus behaving as FP receptor agonists.

The Hydrolysis of Bimatoprost in Corneal Tissue Generates a Potent Prostanoid FP Receptor Agonist

Using human and bovine corneal tissue, we investigated the in vitro metabolism of bimatoprost (17-phenyl-18,19,20-trinor-prostaglandin F 2α ethyl amide, Lumigan (Allergan, Inc, Irvine, CA). Enzymatic amidase activity, which converts bimatoprost to the corresponding prostaglandin carboxylic acid, was found to be present in corneal tissue from both species. Using HPLC and mass spectrometry for analyses, conversion of bimatoprost to 17-phenyl-18,19,20-trinor prostaglandin F 2α continued for at least 24 hours after excision of the cornea, with a conversion rate of approximately 25 μg/24 hours. This hydrolysis product is identical to the free acid of latanoprost with the exception of a double, rather than a single, bond at the carbon 13-14 position. Assuming that this conversion also occurs in vivo at a similar rate, this hydrolysis product may account for the reduction of intraocular pressure occurring in patients treated with bimatoprost.

Synthesis and structure-affinity-activity relationships of novel benzofuran derivatives as MT(2) melatonin receptor selective ligands.

A series of N-(2-phenylbenzofuran-3-yl) ethyl amide and N-(2-arylalkylbenzofuran-3-yl) ethyl amide derivatives were synthesized and evaluated as melatonin receptor ligands. The affinity of each compound for the two MT(1) and MT(2) melatonin receptor subtypes was determined by binding studies using 2-[(125)I]iodomelatonin on human embryonic kidney cell line HEK293 membrane homogenates. The intrinsic activity of the most interesting compounds was evaluated on the [(35)S]GTPgammaS binding assay. Introduction of a 2-phenyl substituent in the C-2 benzofuran position leads to an agonist compound, 10q, which binds more strongly than melatonin itself to both MT(1) and MT(2) subtypes. On the other hand, a 2-benzyl group in the same position allows MT(2) antagonist selective ligands to be obtained. The MT(2) selectivity and antagonist potency can be modulated with suitable modifications on the N-acyl and benzyl substituents, and the most selective compounds 10c and 19 show affinity ratios of 123 and 192, respectively, and bind to the MT(2) subtype similarly to melatonin itself (0.1 nM). Nevertheless, 10c acts as an MT(1) and MT(2) antagonist, whereas 19 is a partial agonist.

Bimatoprost and its free acid are prostaglandin FP receptor agonists

Bimatoprost (17-phenyl-prostaglandin F 2α ethyl amide) has been reported not to exert its actions via prostaglandin receptors. Here, bimatoprost displaced [ 3H]prostaglandin F 2α from FP receptors ( K i=6310±1650 nM). Bimatoprost rapidly mobilized intracellular Ca 2+ ([Ca 2+] i) via cloned human FP receptors expressed in human embryonic kidney cells (EC 50=2940±1663 nM) and via native FP receptors in 3T3 mouse fibroblasts (EC 50=2200±670 nM). Furthermore, AL-8810 ((5 Z, 13 E)-(9 S,11 S,15 R)-9,15-dihydroxy-11-fluoro-15-(2-indanyl)-16,17,18,19,20-pentanor-5,13-prostadienoic acid), an FP receptor antagonist, blocked the bimatoprost-induced [Ca 2+] i mobilization.

Synthesis and anticonvulsant activity of N,N-phthaloyl derivatives of central nervous system inhibitory amino acids

In order to study the influence of the length of the amino acid chain of N,N-phthaloylamino acid amides as analogues of the former anticonvulsant taltrimide on the seizureantagonizing activity glycine, β-alanine and γ-aminobutyric acid (GABA) derivatives were synthesized. The corresponding taurine derivatives were also included. Generally, the glycine-derived amides showed a higher activity than the β-alanine and GABA derivatives in the maximal electroshock seizure (MES) test in mice upon intraperitoneal administration. The activity was comparable to the respective taurine derivatives. The N,N-phthaloyl-glycine amides were also active in the MES test upon oral administration to rats. No significant activity was noted in the seizure threshold test with subcutaneous pentylenetetrazole. The ED50 of N,N-phthaloyl-glycine ethyl amide (4b) in the MES test upon intraperitoneal administration to mice was 19.1 mg/kg. On a molar basis this activity is comparable to the activity of phenytoin with little toxicity in the rotorod test. In conclusion, N,N-phthaloyl-glycine amides might represent promising antiepileptic drugs.

Synthesis and anticonvulsant activity of N,N-phthaloyl derivatives of central nervous system inhibitory amino acids.

In order to study the influence of the length of the amino acid chain of N,N-phthaloyl-amino acid amides as analogues of the former anticonvulsant taltrimide on the seizure-antagonizing activity glycine, beta-alanine and gamma-aminobutyric acid (GABA) derivatives were synthesized. The corresponding taurine derivatives were also included. Generally, the glycine-derived amides showed a higher activity than the beta-alanine and GABA derivatives in the maximal electroshock seizure (MES) test in mice upon intraperitoneal administration. The activity was comparable to the respective taurine derivatives. The N,N-phthaloyl-glycine amides were also active in the MES test upon oral administration to rats. No significant activity was noted in the seizure threshold test with subcutaneous pentylene-tetrazole. The ED50 of N,N-phthaloyl-glycine ethyl amide (4b) in the MES test upon intraperitoneal administration to mice was 19.1 mg/kg. On a molar basis this activity is comparable to the activity of phenytoin with little toxicity in the rotorod test. In conclusion, N,N-phthaloyl-glycine amides might represent promising antiepileptic drugs.

Bombesin-like peptides depolarize rat hippocampal interneurones through interaction with subtype 2 bombesin receptors.

1. Whole-cell patch-clamp recordings were made from visually identified hippocampal interneurones in slices of rat brain tissue in vitro. Bath application of the bombesin-like neuropeptides gastrin-releasing peptide (GRP) or neuromedin B (NMB) produced a large membrane depolarization that was blocked by pre-incubation with the subtype 2 bombesin (BB2) receptor antagonist [D-Phe6, Des-Met14]bombesin-(6-14)ethyl amide. 2. The inward current elicited by NMB or GRP was unaffected by K+ channel blockade with external Ba2+ or by replacement of potassium gluconate in the electrode solution with caesium acetate. 3. Replacement of external NaCl with Tris-HCl significantly reduced the magnitude of the GRP-induced current at -60 mV. In contrast, replacement of external NaCl with LiCl had no effect on the magnitude of this current. 4. Photorelease of caged GTPgammaS inside neurones irreversibly potentiated the GRP-induced current at -60 mV. Similarly, bath application of the phospholipase C (PLC) inhibitor U-73122 significantly reduced the size of the inward current induced by GRP. 5. Reverse transcription followed by the polymerase chain reaction using cytoplasm from single hippocampal interneurones demonstrated the expression of BB2 receptor mRNA together with glutamate decarboxylase (GAD67). 6. Although bath application of GRP or NMB had little or no effect on the resting membrane properties of CA1 pyramidal cells per se, these neuropeptides produced a dramatic increase in the number and amplitude of miniature inhibitory postsynaptic currents in these cells in a TTX-sensitive manner.

A novel and efficient method for synthetic carbohydrate conjugate vaccine preparation: synthesis of sialyl Tn-KLH conjugate using a 4-(4-N-maleimidomethyl) cyclohexane-1-carboxyl hydrazide (MMCCH) linker arm.

STn (NeuAcalpha2 --> 6GalNAc alpha-O-Ser/Thr) is a carbohydrate epitope overexpressed in various human carcinomas. Clinical trials are underway using synthetic STn or STn trimeric glycopeptides [STn, cluster; STn(c)] conjugated with keyhole limpet hemocyanin (KLH) as active specific immunotherapy for these cancers. These vaccines have been prepared by conjugating a crotyl ethyl amide derivative of STn or STn(c) to KLH by direct reductive amination after ozonolysis. In the case of STn(c) the conjugation efficiency and the resulting epitope ratios were low. This may be due to steric hinderance of the short spacer arm. To overcome these difficulties, without resynthesis, the STn(c) glycopeptide was modified by attachment of an MMCCH (4-(4-N-maieimidomethyl) cyclohexane-1-carboxyl hydrazide) spacer arm to the aldehyde derivative, and then conjugated with thiolated KLH. This method gave a higher epitope ratio and yield than the direct method. The STn(c)-MMCCH-KLH conjugate induced high titer antibodies in mice against STn(c). This method may be generally applicable for large synthetic oligosaccharides.

Bombesin Receptor Antagonists Block the Effects of Exogenous Bombesin but Not of Nutrients on Food Intake

The endogenous, meal-contingent release of bombesin (BN)-like peptides is thought to contribute to the termination of a meal. In the following experiments the potency of BN receptor antagonists to attenuate the ability of nutrients to suppress food intake was tested. First, the effectiveness of BN receptor subtype antagonists was verified by testing their ability to block the effects of exogenous BN on food intake. Rats were administered intraperitoneal (i.p.) injections of either saline or 0.1 mg/kg [ D-Phe 12,Leu 14]BN (binds both GRP and NMB receptors), [ D-Phe 6]BN(6–13) ethyl amide (binds GRP > NMB), and cyclo-SS-octa (BIM-23042; binds NMB > GRP). Five minutes later rats were administered 8 μg/kg BN (i.p.) and milk intake was measured. Injections of [ D-Phe 12,Leu 14]BN and [ D-Phe 6]BN(6–13) ethyl amide reliably attenuated the ability of BN to suppress milk intake whereas BIM-23042 was ineffective. The results show that the antagonists were behaviorally effective and that exogenous BN may exert its effects on food intake primarily through the GRP receptor subtype. Next, the antagonists were administered either 5 min prior to or 5 min after an intragastric nutrient load or no load in both overnight-deprived and nondeprived rats, and milk intake was then measured. Stomach loads reduced intake and this effect was not attenuated by BN receptor antagonists. Finally, rats were allowed to prefeed and the milk was then removed. Rats were then administered a BN receptor antagonist (0.1 and 1.0 mg/kg) or saline either immediately after the prefeed, 10 min later, or 20 min later. Milk diet was then returned and intake was measured. Peripheral injections of the BN receptor antagonist had no effect compared to saline on milk intake. Collectively, the results indicate that the blockade of peripheral BN peptide receptors is not sufficient to attenuate the satiety signals generated by stomach loads or prefeeding.

Induction of free radicals in hepatocytes, mitochondria and microsomes of rats by ochratoxin A and its analogs

Oxidative damage may be one of the manifestations of cellular damage in the toxicity of ochratoxin A (OA). OA; its three natural analogs, OB, OC and Oα; and three synthetic analogs, the ethyl amide of OA (OE-OA), O-methylated OA (OM-OA), and the lactone-opened OA (OP-OA) were used to study free radical generation in hepatocytes, mitochondria and microsomes from rats. Electron paramagnetic resonance spectroscopy (EPR) using α-(4-pyridyl-1-oxide)- N- tert-butyl nitrone (4-POBN) as a spin trapping agent showed an enhanced free radical generation due to the addition of NADPH to the microsomes. An EPR signal was not observed in the mitochondria and hepatocyte samples when they were treated with a variety of agents. Addition of OM-OA together with NADPH and Fe 3+ to the microsomes resulted in a strong EPR signal compared with the other analogs, whereas the signal could be quenched by the addition of catalase. OM-OA does not have a dissociable phenolate group and does not chelate Fe 3+. The spin adduct hyperfine splitting constants indicated the presence of α-hydroxyethyl radicals resulting from generated hydroxyl radicals, which were trapped by 4-POBN. The results also suggested that the production of hydroxyl radicals by OA does not require a dissociable phenolate group or the prior formation of an OA-Fe complex.

Effects of bombesin on behavioral thermoregulation in the bullfrog.

Bombesin is a member of a class of neuroactive chemicals that have potent thermoregulatory effects in ectothermic and endothermic vertebrate species. Bombesin-like peptides are found in the brains of ectothermic and endothermic vertebrates and have been implicated in the central nervous system modulation of behavioral thermoregulation. Amphibians rely on behavioral thermoregulation to maintain their body temperature within developmental stage-dependent critical limits. To investigate the influence of bombesin on behavioral thermoregulation, we examined the effects of central injections of bombesin on thermal habitat selection at different stages of bullfrog development. Tadpoles and adult male and female frogs were allowed to select a preferred temperature, within an aquatic thermal gradient, before and after receiving an intracerebroventricular injection of bombesin. In larval and adult female bullfrogs, bombesin administration caused a decrease in preferred temperature values. This effect was clearly dose-dependent in tadpoles. Bombesin effects were variable in adult males, probably due to an overriding stress response to handling exhibited by males. The bombesin-induced hypothermia was blocked by [D-Phe6, Des-Met14]-bombesin (6-14), ethyl amide, a bombesin/gastrin-releasing peptide receptor antagonist. These data suggest that bombesin/gastrin-releasing peptide receptors are functional in the central nervous system of larval and adult amphibians and that receptor binding can modulate thermoregulation. They raise the question: under what natural conditions is endogenous bombesin/gastrin-releasing peptide released in the brain to activate thermoregulatory behavior?

Pharmaco-ontogenic modulation of feeding by oxytocin, bombesin, and their antagonists.

Oxytocin (OX) and bombesin (BN) can both suppress food intake in adult rats. In light of important transient fluctuations in peptide and/or receptor expression early after birth, this study characterized the feeding-suppressant effects of OX and BN during early development and explored their physiological relevance, using BN or OX antagonists, [D-Phe6,des-Met6-14]BN(6-14), ethyl amide (des-Met), and [des-Glycinamide9,d(CH2)5',O-Me-Tyr2,Thr4,Orn8]-vaso toc in (vasotocin), respectively. On postnatal days (PD) 1, 5, 10, and 15, groups of food-deprived Sprague-Dawley rat pups (n = 8-11) were injected SC with saline (control) or BN (0.6, 0.06, or 0.006 mg/kg), des-Met (10 mg/kg), OX (1.2, 0.6, 0.3, or 0.15 mg/kg), or vasotocin (1.0 mg/kg), and their intake of milk (from a piece of absorbent paper saturated with warm milk) was monitored. Results revealed that BN and OX suppressed milk intake from PD 1 to PD 15. Although the milk intake varied with the peptide dose, this effect was age dependent. The doses of BN (but not OX) required to suppress feeding were higher than those needed in adults. When administered alone, OX or BN antagonists did not affect food intake, except at PD 15 for des-Met and PD 1 and PD 10 for vasotocin, where they enhanced feeding. These results suggest that pharmacological effects to OX and BN are apparent from hours after birth and that these peptides may play a role in the regulation of ingestive behavior from early on in ontogeny.

Long-Term Somatotropic and Galactopoietic Effects of a (1-30) Ethyl Amide Analog of Growth Hormone-Releasing Factor

Thirty-five lactating Holstein cows (165±8 DIM) received the following treatments for 96 d: uninfused controls; i.v. infusion of .9, 2.7, or 8.1 mg/d of Ile2, Ser8,28, Ala15, Leu27, Hse30-bovine growth hormone-releasing factor (1–30) ethyl amide; or i.v. infusion of 12 mg/d of recombinant Leu27, Hse45-bovine growth hormone-releasing factor (1–45) lactone. Concentrations of somatotropin in serum and SCM yield of cows infused with 1–30 releasing factor increased quadratically as the dose increased. Responses for somatotropin and SCM yield were quantitatively similar for cows infused with 2.7 and 8.1mg of 1–30 or 12mg of 1–45. While concurrently infused with the 1–30 or 1–45 as described, cows retained their ability to release somatotropin following an acute i.v. injection of 10 nmo1/100kg of BW of either growth hormone-releasing factor 1–30 or 1–45. Mean concentrations of IGF-I in serum increased similarly in magnitude for cows infused at all doses of 1–30 or 1–45. The 1–30 releasing factor generally increased IGF-binding protein-3, but had little effect on IGF-binding protein-2. The 1–45 releasing factor did not significantly affect either binding protein. Yield of SCM was correlated with serum concentrations of somatotropin, but not with IGF-I. Concentrations of NEFA in serum were elevated through 36 and 50 d in response to the highest doses of 1–30 and 1–45, respectively. Treatment did not affect DMI, BW, or body condition score. In conclusion, continuous i.v. infusion of 2.7 and 8.1 mg/d of 1–30 and 12 mg/d of 1–45 similarly increased yield of SCM, somatotropin, and IGF-I without inducing refractoriness in the ability of the anterior pituitary gland to release somatotropin in response to a concurrent, acute challenge with 1–30 or 1–45.