Ontogeny Of Receptors Research Articles

Ontogeny of peripheral-type benzodiazepine receptors in cultured astrocytes and brain from rat

Peripheral-type benzodiazepine receptors (PBRs) in brain are primarily localized within astroglial cells, and the existence of PBR subtypes have been suggested. In the present study the ontogeny of PBRs labeled with [ 3H]Ro5-4864 and [ 3H]PK 11195 in cultured astrocytes was compared to their postnatal in-vivo development. The density of [ 3H]Ro5-4864 binding sites in cultured astrocytes from rat cortex progressively increased from 1- to 3-week-old cultures and did not change further in 5- and 8-week-old cultures. The density of [ 3H]PK 11195 binding sites progressively increased from 1- to 5-week-old cultures. The density of [ 3H]PK 11195 binding sites exceeded the density of [ 3H]Ro5-4864 binding sites by 40–50% in 2-, 5- and 8-week-old cultures. The affinity of the PBR ligands for the receptor sites was increased by 3- to 4-fold from the first to the second week in cultures, and did not change thereafter. A similar developmental pattern of PBRs was observed in rat cortex, except that: first, the difference between the B max of [ 3H]PK 11195 and [ 3H]Ro5-4864 was already apparent in postnatal-week-1 and persisted with maturation; second, the high affinity of the ligands for the receptor sites was apparent from postnatal-week-1 and did not change with maturation. Age-related differences in the ratio between the density of PBRs in astrocytes and rat cortex were also observed. These results lead us to suggest that the development of PBRs in vivo during the first postnatal week is more rapid than the development of the receptors in vitro during the first week in culture. Subsequently, the increased ratio between the density of PBRs in cultured astrocytes and brain with maturation indicates the predominantly astrocytic localization of these receptors. The finding that the density of [ 3H]PK 11195 binding sites in cultured astrocytes and in rat brain cortex is usually 40–50% greater than the density of [ 3H]Ro5-4864 binding sites further supports the existence of PBR subtypes.

Steroid modulation of central nervous GABAA receptor binding in three female rat brain areas during postnatal development.

The ontogenetic development of gamma-aminobutyric acid type A (GABAA) receptors and their sensitivity to neuroactive steroid agonists were determined between birth and day 25 postnatally, using membrane fractions of distinct rat brain areas. At birth, the medulla oblongata (MED) reached adult numbers of GABA-binding sites whose affinity did not change significantly postpartum. In contrast, in the phylogenetically younger frontal cortex (FC) and cerebellum (CER), the number of binding sites increased in a linear fashion, albeit with a greater delay in the CER. Moreover, the affinity varied significantly with age, but in the opposite way. Despite unchanged basal binding in the MED, both the FC and the MED displayed significant increases in the number of GABAA-binding sites elicited by either 3 alpha-hydroxy-5 alpha-pregnane-20-one (THP) or 3 alpha, 21-dihydroxy-5 alpha-pregnane-20-one (THDOC) in vitro (final concentration 1 microM) for all ages investigated, but the magnitude of enhancement was age-dependent. At 15 days postpartum, both THP and THDOC enhanced the density of binding sites in MED with maximal increases by 485 or 462% of the control determined in the absence of steroids. However, the simultaneously observed decrease of affinity did not reach statistical significance. Likewise, in the FC the steroidal influence on affinity did not begin to change until 15 days postpartum. However, there was a significant increase in affinity coincident with a reduced increase in receptor density. The present results demonstrate a brain-area-specific ontogeny of GABAA receptors and developmental changes in the sensitivity of GABAA-binding sites to steroids.

Perinatal ontogeny of porcine nuclear thyroid hormone receptors and its modulation by thyroid status

Induction of nuclear thyroid hormone receptors (TRs) represents a key point in the control of growth, development, differentiation, and metabolism of most tissues. The influence of thyroid status on the ontogeny of hepatic and skeletal muscle TRs has been investigated in perinatal pigs. Plasma concentrations of total and free 3,5,3'-triiodothyronine (T3) increased markedly from 80 days of fetal life (80 f) to 2 days of postnatal life. Test piglets obtained from sows fed a high glucosinolate rapeseed diet had lower T3 and thyroxine levels than controls at 110 f and showed a higher postnatal surge in T3. Maximal T3 binding capacity (Bmax, pmol T3/mg DNA, means +/- SE) in liver increased from 0.07 +/- 0.01 at 80 f to 0.37 +/- 0.02 at birth and then plateaued. In longissimus dorsi muscle, Bmax values were much higher than in liver and increased from 0.90 +/- 0.02 at 80 f to 1.37 +/- 0.13 at birth and then declined to 1.09 +/- 0.11 at 2 days of age. Long-term fetal hypothyroidism affected the ontogenic profile of both liver and muscle receptors but in opposite directions; Bmax values were reduced in liver but increased in muscle. Postnatally, lower muscle Bmax values occurred in parallel with transient higher levels of circulating T3. Apparent binding affinities were slightly different in liver and muscle during fetal life, and there was an effect of age in muscle. In conclusion, as far as the receptor is concerned, fetal muscle can potentially respond to thyroid hormones much earlier in development than the liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of in utero ethanol exposure on the postnatal ontogeny of insulin-like growth factor-1, and type-1 and type-2 insulin-like growth factor receptors in the rat brain

There is convincing evidence that alcohol consumption during pregnancy causes major CNS abnormalities; however, the molecular and cellular basis of these dysfunctions is currently not understood. This study examined the effects of prenatal ethanol exposure on the expression of insulin-like growth factor-1 messenger RNA and type-1 and type-2 receptor protein and messenger RNA expression in the developing rat brain. Mothers were maintained on an ethanol containing liquid diet from day 2 of pregnancy through parturition and the offspring were killed at birth, 10, 20 and 40 days of age. Insulin-like growth factor-1 messenger RNA, and insulin-like growth factor receptors demonstrated developmentally dependent expression in specific brain regions throughout the postnatal period of CNS maturation. Insulin-like growth factor-1 gene expression in the brain, as analysed by dot-blot hybridization, was greatest at birth, and decreased 61% in ad libitum and pair-fed animals by 20 days of age. In contrast, ethanol-treated animals exhibited only a 25% decrease in insulin-like growth factor-1 messenger RNA levels during the same period. This delay in insulin-like growth factor-1 messenger RNA maturation may be related to a developmental delay in CNS development in the prenatally ethanol exposed offspring. Prenatal ethanol exposure did not alter the observed localization of insulin-like growth factor-1 messenger RNA. While alterations were observed in long-term insulin-like growth factor-1 messenger RNA regulation, quantitative receptor autoradiography and in situ hybridization demonstrated no alterations in either type-1 or type-2 insulin-like growth factor receptor populations in ethanol-treated animals. Changes in hepatic and plasma insulin-like growth factor-1 and insulin-like growth factor-binding protein regulation have also been observed in these animals, suggesting changes in protein translation and the autocrine/paracrine actions of this peptide. The present study demonstrated that insulin-like growth factor-1 messenger RNA and insulin-like growth factor receptors are regionally expressed during early postnatal development and that ethanol administration influenced the long-term regulation of insulin-like growth factor messenger RNA levels in the brain without affecting either its localization or insulin-like growth factor receptor populations. These results, in combination with previous results from our laboratory on the effects of prenatal ethanol exposure on plasma insulin-like growth factor-1 and insulin-like growth factor-binding proteins, provide additional evidence that even modest levels of prenatal ethanol exposure enhance the possibility of alterations in growth promoting peptides in the fetus and neonate, leading to an increased risk of fetal alcohol effects in the offspring.

D 1- and D 2-receptor mediation of motoric behavior in rats depleted of DA as neonates: Effects of age and size of depletion

The D 1/D 2 mediation of motor behavior in rats is qualitatively altered following large forebrain dopamine (DA) depletions on postnatal day 3. These animals are markedly subsensitive, relative to controls or animals depleted of DA as adults, to the motoric deficits produced by individual D 1- or D 2-like antagonists but are impaired following combined D 1 + D 2 antagonists. In order to determine the extent of DA depletion necessary to produce this subsensitivity to individual antagonists, we compared the motoric effects of D 1 and D 2 antagonists in adult animals sustaining a wide range of DA depletions on day 3. Only animals with striatal depletions of ≥95% demonstrated this subsensitivity to individual antagonists. Moreover, since important changes in DA receptor ontogeny occur during the first postnatal week, we compared the ability of the D 1-like antagonist, SCH 23390 (0.2 mg/kg), or the D 2-like antagonists, clebopride (10.0 mg/kg) and haloperidol (1.0 mg/kg), to induce akinesia or catalepsy in adults depleted of DA on either day 1 or on day 3. Rats depleted of striatal DA (>95%) at either age exhibited similar subsensitivity to D 1 or D 2 antagonists. These findings suggest that large DA depletions are necessary to alter the roles of D 1 and D 2 receptors in the expression of motor behavior and that this plasticity is comparable in animals depleted on day 1 or day 3.

In search of a mechanism for receptor-mediated neurobehavioral teratogenesis by nicotine: catecholamine release by nicotine in immature rat brain regions

Nicotine disrupts central nervous system development through interactions with nicotinic cholinergic receptors found in immature brain, leading to discoordination of target cell replication and differentiation. However, it is unclear whether the net result is achieved by nicotine's actions on its specific target cells, or indirectly through receptor-mediated release of other neurotransmitters, such as catecholamines, that possess neurotrophic properties. In the current study, developing rats (1, 7, 14 and 21 days old) were challenged acutely with nicotine (0.3 mg/kg) and the release of catecholamines was evaluated in vivo (AMPT method) in three brain regions that differ in nicotinic receptor concentrations. Nicotine did not stimulate catecholamine release at birth, but developed the capacity to do so in parallel with the ontogeny of nicotinic cholinergic receptors in the midbrain + brainstem and in the forebrain. In the cerebellum, which remains poor in nicotinic receptors, no response was obtained at any age. Superimposed on this general pattern, changes in sensitivity to nicotine were also seen that corresponded to ontogenetic changes in endogenous cholinergic tone, suggesting that receptor desensitization occurs normally during developmental stages in which neuronal activity is high. The absence of a catecholamine response to nicotine at birth in the rat indicates that neurobehavioral teratology associated with fetal nicotine exposure does not reflect secondary actions mediated through catecholamines. However, because brain development in the neonatal rat corresponds to fetal stages in man, the onset of these mechanisms may be relevant to human fetal exposure.

Ontogeny of glucocorticoid and D2 receptors in the rat pituitary: an in situ hybridization study

The expression of glucocorticoid and D2 dopamine receptors (GR and D2R) during rat pituitary ontogenesis was studied by in situ hybridization (ISH). On early stages, E13–E14, a weak specific signal for GR mRNA was obvious in the whole Rathke's pouch (RP) whereas subsequently, from E17–E18, strong labelling was restricted to the anterior lobe (AL) and the neural lobe (NL). At the same time, D2R mRNAs appeared in the intermediate lobe (IL) and the long isoform of the D2R (D2R 444) was detectable with specific probes. On the postnatal stages, until adult, GR mRNA, if present, was always undetectable in the IL using the conventional ISH technique. These data indicate a possible early regulation of POMC gene expression by glucocorticoid in corticotrophic cells of the AL and by dopamine in the melanotrophic cells of the IL. The possibility of a negative regulation of GR mRNA by dopamine (DA) in the IL as soon as E17 is discussed.

Ontogeny of neuropeptide FF pharmacology and receptors in mouse brain

The postnatal ontogeny of neuropeptide FF (NPFF) receptors in mouse brain was mapped by quantitative autoradiography using [ 125I](1DME)Y8Famide as selective radioligand. By day 14, the adult distribution of NPFF sites was established in almost all brain areas. The density of NPFF receptors reached adult levels between 14 and 28 days after birth, earlier than did μ or δ opioid receptors. During ontogeny, morphine analgesia was measured in hindpaw, forepaw and tail immersion tests (57°C). An antinociceptive effect of morphine in the forepaw and tail immersion test was evident in 14-day-old but not in 7-day-old mice. In 14 and 21 day-old mice, the influence of (1DME)Y8Famide on morphine analgesia depended on the dose and on the particular test. A low dose of (1DME)Y8Famide inhibited morphine analgesia in the tail or hindpaw test, whereas a higher dose had either no effect (tail test) or increased morphine activity (hindpaw test). In adult mice, only the inhibition of morphine antinociception by (1DME)Y8Famide was observed. These observations revealed the establishment of an equilibrium between NPFF and opioid functions during ontogeny.

Glucocorticoid Receptor mRNA Ontogeny in the Fetal and Postnatal Rat Forebrain

Glucocorticoid receptor (GR) ontogeny and distribution in postnatal rat brain have been demonstrated, but onset and distribution of GR gene expression during fetal life has not been reported. This study focuses on the distribution of GR-mRNA in the fetal and postnatal rat forebrain, with emphasis on hypothalamic and limbic structures. Time pregnant rats were decapitated at 8:309:30 AM on Gestational Days 14 (F14), F16, F17, F18, and F19. Postnatally, rats were sacrificed on Days 1, 4, 6, 10, and 16. Cryostat sections were subjected to in situ hybridization, using a cRNA probe directed to the GRmRNA. GR-mRNA was detectible in the hippocamposeptal formation as early as F14, By F16, GR gene expression was evident in the hypothalamic paraventricular nucleus (PVN) as well. During late gestation (F17-F19), GR-mRNA was localized also in the thalamus, hippocampus, amygdala, and discrete cortical regions. Postnatally, GR-mRNA abundance was high in the PVN, CA1/CA2 hippocampal field, piriform cortex and dorsal endopiriform nucleus, specific amygdaloid nuclei, and the suprachiasmatic nucleus. In PVN, GR-mRNA was present prior to the onset of CRH gene expression (F17), which may suggest a role for GR in neuronal differentiation.

Ontogeny of vasoactive intestinal peptide receptors in rat ventral prostate

1. 1. The stimulatory effect of VIP on rat prostatic adenylyl cyclase changes during postnatal development. It peaks at 2 months (peripubertal period), remains in a plateau between 3 and 12 months (adult period), and decreases at 24 months (old period). 2. 2. The stimulation of rat prostatic adenylyl cyclase by the β-adrenergic agonist isoproterenol follows a pattern rather similar to that of VIP. 3. 3. The values of VIP binding capacity correlate well with those observed for adenylyl cyclase between 1 and 12 months, whereas there appears to exist a great number of uncoupled VIP receptors at 0.5 and 24 months. 4. 4. The apparent molecular mass (51 kDa) of the rat prostatic VIP-receptor complex remains unaltered during ontogenic development.

Functional Ontogeny of Pulmonary Vascular DA1 Dopamine Receptors in the Isolated Perfused Rabbit Lung

Using an in situ isolated salt-perfused rabbit lung preparation, we investigated the functional ontogeny of pulmonary vascular dopamine receptors. In rabbits from 1 to 23 d of age, we measured pulmonary vascular vasodilatory responses to the peripheral vascular dopamine receptor (DA1) agonist, fenoldopam, and sodium nitroprusside during prostaglandin F2 alpha-induced pulmonary vasoconstriction. In separate experiments, the lungs were pretreated with the DA1 receptor blocker, SCH 23390, before prostaglandin F2 alpha, fenoldopam, and sodium nitroprusside. Lungs from rabbits at one of 6 age groups (n = 6-8 per group) were ventilated and perfused. After a stabilization period, prostaglandin F2 alpha was infused into the pulmonary inflow catheter in a concentration range to yield a sustained rise in mean pulmonary artery pressure (4.9 +/- 0.2 mm Hg). Fenoldopam was injected into the pulmonary artery at doses of 0.01, 0.1, 1.0, and 10 micrograms/g after a recovery period, sodium nitroprusside (0.2 micrograms/g) was injected into the pulmonary artery, and the resultant changes in vascular pressure were recorded. Across all age groups, with and without DA1 receptor blockade, sodium nitroprusside-induced vasodilation was similar (-2.7 +/- 0.2 mm Hg) and was considered reference vasodilation. The fenoldopam vasodilation response was considered a percentage of the sodium nitroprusside reference. Response to fenoldopam varied significantly (p < 0.05 by analysis of variance) across the six age groups, with a maximum at 3-5 d of age. Pretreatment with SCH 23390, a selective DA1-blocking agent, significantly attenuated fenoldopam vasodilation in all but the youngest animals (age 0-2 d), in which no blockade effect was noted.(ABSTRACT TRUNCATED AT 250 WORDS)

Lactoferrin receptors in piglet small intestine: Lactoferrin binding properties, ontogeny, and regional distribution in the gastrointestinal tract

Bioavailability of iron from human milk is known to be high. This may be due to a receptor-mediated mechanism in the small intestine that facilitates the absorption of iron from lactoferrin. Our aim in this study was to evaluate the ontogeny and localization of lactoferrin receptors within the small intestine. The piglet was used as an animal model because lactoferrin is a major iron-binding protein in both human and porcine milk. Kinetics of lactoferrin interaction with its receptor and receptor density were determined in relation to the age of the piglet (day 0–21 after birth) and the location (duodenum, jejunum, and ileum) within the small intestine. Specific and saturable binding of 59Fe-labelled pig lactoferrin by brush border membranes purified from piglet intestine was observed. Pig transferrin, human, and bovine lactoferrin did not bind to the porcine lactoferrin receptor. Lactoferrin binding occurred throughout the intestine independent of age of the piglet; receptor number (15 × 10 14/mg protein) and affinity ( K d = 3 × 10−7 M) were relatively constant from birth until weaning. Thus, it is possible that lactoferrin receptors throughout the intestine may play a role in iron absorption throughout infancy.

The ontogeny of estrogen receptors in heterochronic hippocampal and neocortical transplants demonstrates an intrinsic developmental program

We investigated the intrinsic vs. environmental regulation of estrogen receptor (ER) ontogeny in the neocortex, hippocampus and hypothalamus by employing a heterochronic transplantation paradigm. These studies were based on previous reports demonstrating that neural ER develop asynchronously with quantitatively distinct ontogenetic profiles in various brain regions. Fetal (E14–15) hippocampal, frontal cortical or hypothalamic preoptic area (HPOA) primordial tissue was grafted into frontal cortical lesion cavities made in newborn (PND-0) rats. Thus, the grafted tissue was 1 week younger than the host. Two and 4 weeks following transplantation surgery, which corresponds to a theoretical donor age of PND-7 and PND-21, the grafts, a region of the host neocortex surrounding the transplant, and the host hippocampus, frontal cortex or HPOA (depending on graft type) were assayed for ER content using in vitro binding assays. ER concentration in hippocampal grafts at theoretical age PND-7 were significantly higher than those found in the host (PND-14) hippocampus and in the host neocortex adjacent to the transplant. By theoretical graft age PND-21, ER concentration in hippocampal transplants had decreased to levels comparable to those found in the host. This developmental pattern is analogous to that previously reported for the in situ hippocampus. A similar profile of ER concentration corresponding to the donor age developmental timetable was observed in neocortical grafts. ER levels in HPOA grafts did not change from theoretical donor age PND-7 to PND-21, which also corresponds to the normal ontogenetic profile. These data suggest that region-specific developmental patterns of ER expression in the rat brain are specified by embryonic day 14.

Postnatal ontogeny of GABAA and benzodiazepine receptors in individual layers of rat visual cortex and the effect of visual deprivation.

The postnatal development of GABAA and benzodiazepine receptors was studied in the rat visual cortex with [3H]muscimol and [3H]flunitrazepam, respectively. The studies were performed by quantitative receptor autoradiography in the different cortical layers from normally raised rats and from monocularly deprived animals. In all cortical layers of normal rats there was a sharp (approx 3-fold) increase in [3H]muscimol binding from days 1 to 10. Between postnatal days 10 and 20 there was a considerable decrease in GABAA receptors; thereafter this decline continued until adulthood but at a more moderate rate. In all cortical layers, [3H]flunitrazepam binding increased sharply, also from day 1 to 10; at this age the adult values were practically attained. Monocular deprivation, by unilateral eyelid suture at the age of 11 days onwards lead to an increase in [3H]muscimol binding in cortical layers IV-VI. This increase in GABAA receptors was detectable at the age of 25 days onwards. In contrast, monocular deprivation did not alter the density of benzodiazepine receptors in any of the cortical layers, and this lack of effects was regardless of the duration of visual deprivation. The present results suggest that the development of GABAA receptors in the deeper cortical layers, but not that of benzodiazepine receptors, is seemingly regulated by the visual experience at a critical period of brain maturation.

Ontogeny of corticosteroid receptors in the brain.

1. In the brain, glucocorticoids bind to both the glucocorticoid receptor (GR) and the mineralocorticoid receptor (MR). These receptors show clearly distinct developmental patterns in the infant rat. 2. Low levels of GRs are present around the time of birth throughout the brain. Concentrations rise slowly, and do not achieve adult levels until the third week of life, approximately. GR affinity for corticosterone is higher perinatally than at later ages. Receptor microdistribution changes dramatically during ontogeny. In particular, certain regions, such as the suprachiasmatic nucleus of the hypothalamus, express high levels of receptor only during the first week of life. GRs may show impaired capacity to undergo transformation and/or nuclear translocation during the second postnatal week. Environmental manipulations during early ontogeny (e.g., early handling) may have permanent effects on GR capacity. 3. MRs are present at very low concentrations in the first days of life. Binding capacity rises rapidly thereafter and resembles that found in the adult by the end of 1 week. Neither binding affinity in vitro nor overall distribution changes with age. As in the adult, low doses of corticosterone, in vivo, bind mainly to the MRs. Levels of corticosterone are low and relatively unperturbable in the intact infant rat. It is likely, therefore, that most of the physiological actions of this hormone during this period are mediated by the MR.

Ontogeny of thyroid hormone receptors and c-erbA expression during brown adipose tissue development: evidence of fetal acquisition of the mature thyroid status

We have determined the developmental changes in thyroid hormone receptor expression and thyroid hormone status in rat brown adipose tissue (BAT). The present study demonstrates that c-erbA alpha and -beta genes are expressed in the developing BAT. The 6.5-kilobase (kb) beta 1, 2.6-kb alpha 2, 5.5-kb alpha 1, and 6.6-kb alpha transcripts showed peak values on day 20 of fetal life. High affinity T3-binding sites were found in fetal BAT. Binding capacity was similar (18-day-old fetuses) or higher (20-day-old fetuses) than in postnatal or adult brown fat. In contrast, T3 receptors were scarce in fetal liver, and values in adult liver were similar to those in fetal BAT on day 20. The profiles of iodothyronine 5'-deiodinase, nuclear T3 content, and receptor occupancy also showed peak values in fetal BAT on day 20, but they were very low in fetal liver. Thus, BAT has already achieved complete maturation of its thyroid status on day 20 of gestational age. This highly tissue-specific developmental pattern is unique among other mammalian thyroid-sensitive tissues studied to date. Between days 18 and 20 of rat fetal development, there is a specific induction of the uncoupling protein gene expression, the main marker of differentiated adipocytes. The results suggest that thyroid hormones may be involved in the establishment of the differentiated phenotype of the brown fat cell in fetal life.

Testicular descent. II. Ontogeny and response to denervation of calcitonin gene-related peptide receptors in neonatal rat gubernaculum

The gubernaculum guides inguino-scrotal testicular descent by migrating into the scrotum ahead of the testis. Calcitonin gene-related peptide (CGRP) is present in the genitofemoral nerve of the neonatal rat and stimulates gubernacular motility in vitro. In a previous study in vitro autoradiography demonstrated a distinctive distribution of binding sites for CGRP over the developing cremasteric muscle in the gubernaculum. Binding analysis using computerized densitometry revealed a single class of sites. This study aimed to characterize the ontogeny of CGRP receptors in the gubernaculum and their response to denervation. Gubernacular sections from neonatal male rats were incubated with [125I]human CGRP as well as a variety of unlabeled neuropeptides. The expression of CGRP receptors culminates during the first week after birth, when gubernacular migration actually occurs. Significantly higher binding capacities were found in the denervated gubernacula compared with those in controls, which suggests an upregulation of CGRP receptors as a result of the genitofemoral nerve denervation. These results are consistent with the hypothesis that CGRP released from the nerve acts directly on the developing cremaster via its own receptors, which have not been described previously in this tissue.

Testicular descent. II. Ontogeny and response to denervation of calcitonin gene-related peptide receptors in neonatal rat gubernaculum.

The gubernaculum guides inguino-scrotal testicular descent by migrating into the scrotum ahead of the testis. Calcitonin gene-related peptide (CGRP) is present in the genitofemoral nerve of the neonatal rat and stimulates gubernacular motility in vitro. In a previous study in vitro autoradiography demonstrated a distinctive distribution of binding sites for CGRP over the developing cremasteric muscle in the gubernaculum. Binding analysis using computerized densitometry revealed a single class of sites. This study aimed to characterize the ontogeny of CGRP receptors in the gubernaculum and their response to denervation. Gubernacular sections from neonatal male rats were incubated with [125I]human CGRP as well as a variety of unlabeled neuropeptides. The expression of CGRP receptors culminates during the first week after birth, when gubernacular migration actually occurs. Significantly higher binding capacities were found in the denervated gubernacula compared with those in controls, which suggests an upregulation of CGRP receptors as a result of the genitofemoral nerve denervation. These results are consistent with the hypothesis that CGRP released from the nerve acts directly on the developing cremaster via its own receptors, which have not been described previously in this tissue.

Ontogeny and characterization of epidermal growth factor receptors on the fetal area of the sheep placenta

Studies of the binding of 125I-labelled epidermal growth factor (EGF) to the ovine placenta were carried out on days 50, 90-100 and 140 of pregnancy. Membrane fractions were purified from the fetal area of the cotyledon. Two classes of binding sites were found. Their dissociation constants (Kd) were not significantly different for the three stages of pregnancy considered (high-affinity Kd 54-70.2 pmol/l; low-affinity Kd 12.2 to 19 nmol/l). However, the number of high-affinity binding sites on days 90-100 was significantly (P < 0.01) greater (146 +/- 19 fmol/mg protein) than on either day 50 or day 140 (respectively 74.2 +/- 1.26 and 56.3 +/- 5.6 fmol/mg protein). Affinity cross-linking studies followed by SDS-PAGE under reducing conditions demonstrated that the major part of the EGF was specifically cross-linked to a protein of molecular weight of 150 kDa and to lesser extent to 180 kDa and 130 kDa proteins. Membranes prepared from unfrozen tissues, in the presence of sodium iodoacetate to reduce endogenous enzymatic conversion of the 180 kDa form to the 150 and 130 kDa forms, still exhibited a major EGF-binding protein of 150 kDa. The occurrence of an increased number of EGF receptors at the period of rapid cotyledonary growth which coincides with the increase in placental hormonal secretions suggests that EGF has a role in the development of the ovine placenta.

Postnatal laminar development of cholinergic receptors, protein kinase C and dihydropyridine-sensitive calcium antagonist binding in rat visual cortex. Effect of visual deprivation

The postnatal ontogeny of 3H-pirenzepine and 3H-oxotremorine-M binding to M1- and M2-muscarinic acetylcholine receptors, respectively, as well as 3H-nicotine binding to neuronal nicotinic acetylcholine receptors, 3H-phorbol-12,13-dibutyrate binding to protein kinase C and 3H-PN200-110 binding to dihydropyridine-sensitive calcium channels was studied in individual layers of the visual cortex in both normally raised and monocularly deprived rats (one eyelid sutured at the age of 11 days) using quantitative receptor autoradiography. Postnatal ontogeny of M1-muscarinic receptors is similar in each visual cortical layer reaching the highest receptor density at the age of 15 days, whereas M2-muscarinic binding sites increase gradually from day 7 up to day 34. Highest 3H-nicotine binding is reached in all visual cortical layers at postnatal day 15 followed by a considerable decrease in binding sites until day 25. Phorbol ester binding rises considerably from birth until the age of 15 days reaching nearly the adult value in the upper layers, whereas in layers V and VI a marked decrease in binding levels until adulthood can be observed. The developmental course of 3H-PN200-110 binding sites is similar in all visual cortical layers and exhibits a moderate rise in binding sites between postnatal days 7 and 15. Monocular deprivation results in permanent changes in the developmental profiles of phorbol ester as well as calcium antagonist binding sites, whereas the alterations in muscarinic and nicotinic cholinergic receptors folloing monocular deprivation are only of transient nature. The data presented suggest that acetylcholine plays a modulatory role during a certain period of early postnatal maturation of the visual cortex by affecting both cholinergic receptors and associated second messenger cascades.