Nisar A. Pampori

Differential effects of neonatally administered glutamate on the ultradian pattern of circulating growth hormone regulating expression of sex-dependent forms of cytochrome P450

Neonatal male rats were treated with monosodium glutamate (MSG) at either 0.5, 1.0, 2.0, 3.0 or 4.0 mg/g body weight on alternate days during the first 9 days of life. As adults, rats were catheterized to obtain unstressed, serial blood samples for the determination of ultradian patterns of circulating growth hormone. In addition, the levels of drug-metabolizing enzymes (i.e. hexobarbital hydroxylase, cytochromes P450 and b5, NADPH-cytochrome P450 reductase and ethoxyresorufin O-deethylase) as well as sex-dependent forms of cytochrome P450 [i.e. male-dependent cytochromes P450 2c (IIC11), 2a (IIIA2) and RLM2 (IIA2) and female-dependent cytochromes P450 2d (IIC12) and 3 (IIA1)] and/or their catalytic activities were measured in the hepatic microsomes of the treated rats. The results demonstrated a dose-dependent, graded response to MSG treatment. As the dose of MSG increased from 0.5 to 4.0 mg, there was a concurrent decline in the amplitudes of the characteristically masculine, episodic bursts of growth hormone, until at the highest dose (4 mg), the pulses were no longer detectable. Associated with this dose-dependent alteration in the ultradian pattern of growth hormone secretion was a measurable change in the activities of the sex-dependent hepatic enzymes. As the pulse heights of the hormone declined to 10-20% of their normal amplitudes, the levels of the male-dependent enzymes (i.e. the drug-metabolizing enzymes, as well as the male forms of cytochrome P450 and their specific steroid hydroxylases) were maintained, and in some cases, exceeded the levels normally found in males. However, as the hormone pulse heights declined, there appeared an accompanying increase in the activities of some of the female-dependent enzymes. Finally, with the loss of all detectable levels of circulating growth hormone, the normal masculine profile of hepatic enzymes was reversed to an apparently normal (with the exception of cytochrome P450 2d) feminine profile. Summarizing, the results indicate that (1) neonatal administration of MSG can produce dose-dependent, graded, long-term developmental defects in the ultradian rhythm of circulating growth hormone and associated sex-dependent hepatic enzymes, and (2) while the male-dependent hepatic enzymes can be maintained at normal or even higher levels in the face of an up to 90% reduction in the pulse heights of plasma growth hormone, the activities of the female-dependent enzymes may begin to increase.

Sexual dimorphism in avian hepatic monooxygenases

Adult white Leghorn chickens exhibited a sexual dimorphism in hepatic microsomal monooxygenases determined from the concentrations of total cytochromes P450 and b5, and the metabolism of drug (hexobarbital, coumarin and ethoxyresorufin) and steroid (androstenedione and testosterone) substrates that were 2- to 4-fold greater in roosters than in hens. Caponizing at 6 weeks of age reduced the activities of the monooxygenases to levels comparable to those found in intact hens. In spite of the fact that testosterone replacement maximally stimulated comb growth in the capons and elevated (i.e. masculinized) hepatic monooxygenase activities in the hens to male-like levels, androgen replacement was ineffective in increasing the subnormal enzyme levels in the capons. While the failure of testosterone administration to restore monooxygenase levels in the capons may be explained by the immaturity of the birds at orchiectomy, the present results demonstrate, that like some mammals, birds may display gender differences in hepatic monooxygenases that are regulated by the testes.

Effects of neonatally administered monosodium glutamate on the sexually dimorphic profiles of circulating growth hormone regulating murine hepatic monooxygenases

Neonatal male and female mice were treated with monosodium glutamate (MSG) at either 2.0 or 4.0 mg/g body weight on alternate days during the first 9 days of life. As adults, mice were catheterized to obtain unstressed, serial blood samples for the determination of ultradian profiles of circulating growth hormone. In addition, monooxygenase levels (i.e. steroid hydroxylases and drug-metabolizing enzymes) were measured in hepatic microsomes. Generally, both doses of MSG produced the same developmental defects. Mice neonatally exposed to the amino acid developed a syndrome characterized by retarded growth, obesity and reduced organ weights. While vehicle-treated mice secreted growth hormone in sexually dimorphic patterns defined by pulse frequency (i.e. F > M), hormone concentrations in plasma samples obtained during 8 continuous hr of serial blood collections from both male and female MSG-treated mice were barely detectable at best, and exhibited no pulsatility. Approximately 15% of the measured monooxygenases were male-predominant, 35% were female-predominant and 50% had no sex differences. The enhanced expression of the hepatic monooxygenases in response to MSG-induced depletion of plasma growth hormone indicates that the hormone basically functions as a suppressor of the murine enzyme system.

Subnormal concentrations in the feminine profile of circulating growth hormone enhance expression of female-specific CYP2C12

When newborn female rats were treated with monosodium glutamate, 4 mg/g body weight, on days 1 and 3 of life, circulating growth hormone concentrations were permanently reduced 75-85% in adulthood, whereas the feminine secretory profile characterized by frequent growth hormone pulses, separated by short-lived, measurable troughs, persisted. Associated with this reduction in growth hormone secretion was a mild obesity and a slight depression in peripubertal body weight. In contrast, expression of growth hormone-dependent, female-specific CYP2C12 was increased by almost 100% when measured at both its protein and mRNA levels. In agreement, this supraphysiological expression of CYP2C12 was reflected at a pharmacologic level by a simultaneous elevation in in vitro and in vivo hexobarbital metabolism. When growth hormone secretion was pulsatile (i.e. masculine) or was eliminated from the circulation (i.e. hypophysectomy), hepatic CYP2C12 protein and mRNA were undetectable. The present findings suggest that the normal levels of plasma growth hormone found in female rats are not necessarily optimum for the expression of female-specific CYP2C12.

Nominal growth hormone pulses in otherwise normal masculine plasma profiles induce intron retention of overexpressed hepatic CYP2C11 with associated nuclear splicing deficiency.

Restoration of circulating masculine GH profiles at minipulse amplitudes (i.e. ∼10% of normal) to hypophysectomized male rats and neonatal administration of monosodium glutamate (MSG), producing a similar plasma GH profile, both result in an overexpression (∼200–300%) of CYP2C11 messenger RNA (mRNA), the predominant hepatic cytochrome P450 (CYP) drug-metabolizing enzyme in adult male rats. Coincident with the severalfold elevation in transcript level is a modest 10–30% overexpression of CYP2C11 protein and its catalytic activities. Using hepatic tissue from adult, neonatally MSG-treated rats, we have cloned a variant species of CYP2C11 mRNA containing all of the essential elements of a full-length complementary DNA, including initiating codon, termination codon, and polyadenylase tail. In addition, the transcript contains a 742-bp intervening sequence (identical to the complete terminal intron) between the last and penultimate exons, and an intron-specific oligo probe for Northern blotting demonstrates th...

Sex- and dose-dependent effects of neonatally administered aspartate on the ultradian patterns of circulating growth hormone regulating hexobarbital metabolism and action

Rats, neonatally treated with monosodium aspartate (MSA), exhibited developmental defects through adulthood that were characterized by stunted growth, obesity, and reduced size of the liver, kidney, adrenals, and pituitary. Adult male and female rats treated with 4 mg of MSA had no detectable plasma growth hormone as determined from serial blood samples taken every 15 min for 8 consecutive hr. Associated with this loss of circulating growth hormone in the males was a dramatic decline in in vivo and in vitro hexobarbital metabolism and hepatic cytochrome P450 to female levels. The loss of plasma growth hormone in the females had no effect on the already low levels of hepatic monooxygenases. At 2 mg/g body weight, MSA produced both sex- and dose-dependent effects that were far more subtle than the full-blown obesity and growth retardation associated with the larger 4-mg dose. While the mean concentration of circulating growth hormone was reduced 70 to 90% in 2-mg-MSA-treated rats, the sexually dimorphic, ultradian patterns of growth hormone secretion were undisturbed. Affected males continued to secrete a pulse of growth hormone every 3 hr, albeit at greatly reduced amplitudes, interposed by normally undetectable baselines. Similarly, 2-mg-MSA-treated females had greatly reduced mean levels of plasma growth hormone, but with the usual secretion of multiple pulses never dropping to baseline. Surprisingly, the sex-dependent, hepatic monooxygenases, which are normally regulated by the ultradian secretions of growth hormone, were unaffected by the 2-mg MSA treatment. Our results suggest that while an ultradian pulse of circulating growth hormone is necessary for the characteristically male profile of hepatic monooxygenases, neither the amplitude of the secretory peaks nor their total growth hormone content is critical.