Catestatin and orexin-A neuronal signals alter feeding habits in relation to hibernating states.

Hibernation is a physiological state that by putting vital biological processes at rest enables mammals to protect all organs, especially the brain against ischemic insults and reperfusion injuries. Earlier studies have highlighted the role of hypothalamic (HTH) sites like the periventricular nucleus (Pe) toward sleep-wake and cardiovascular activities of hibernators. In the present work, infusions of Pe with the orexigenic neuropeptide orexin-A (ORX-A) or the novel anti-obesity sympathoinhibitory neuroactive peptide catestatin (CST) have been correlated to differing feeding and motor behaviors in the facultative hibernating hamster Mesocricetus auratus. Behavioral observations showed that treatment with CST provided an anti-obesity activity via the reduction of food intake and body weight for all hibernating states, while ORX-A promoted orexigenic events during mainly the entrance phase. Moreover, hamsters treated with this neuropeptide during the entrance and the arousal hypertensive phases also featured elevated ORX 2 receptor (ORX2R) levels in the third layer of the parietal cortex and lateral HTH (LH), areas involved with feeding, motor plus sleep-wake rhythms. Conversely, ORX-A down-regulated ORX2Rs in the ventromedial (VMH) and supraoptic (SO) HTH nuclei that are associated with anorexigenic effects. Even CST induced mixed ORX2R expression patterns in mostly HTH areas like the evident down-regulation in LH along with the up-regulation in VMH and SO. Overall treatments, especially ORX-A+CST led to reduced neurodegenerative phenomena in HTH supporting their importance together with ORX2Rs in preserving hemodynamic activities, feeding and sleep-wake rhythms of this diencephalic station, which may supply useful therapeutic indications for treating cardiovascular disturbances linked with brain dysfunctions.

Antihypertensive and neuroprotective effects of catestatin in spontaneously hypertensive rats: interaction with GABAergic transmission in amygdala and brainstem.

The chromogranin A-derived peptide catestatin (CST) exerts sympathoexcitatory and hypertensive effects when microinjected into the rostral ventrolateral medulla (RVLM: excitatory output); it exhibits sympathoinhibitory and antihypertensive effects when microinjected into the caudal ventrolateral medulla (CVLM: inhibitory output) of vagotomized normotensive rats. Here, continuous infusion of CST into the central amygdalar nucleus (CeA) of spontaneously hypertensive rats (SHRs) for 15 days resulted in a marked decrease of blood pressure (BP) in 6-month- (by 37 mm Hg) and 9-month- (by 65 mm Hg)old rats. Whole-cell patch-clamp recordings on pyramidal CeA neurons revealed that CST increased both spontaneous inhibitory postsynaptic current (sIPSC) amplitude plus frequency, along with reductions of sIPSC rise time and decay time. Inhibition of GABAA receptors (GABAARs) by bicuculline completely abolished CST-induced sIPSC, corroborating that CST signals occur through this major neuroreceptor complex. Hypertension is a major risk factor for cerebrovascular diseases, leading to vascular dementia and neurodegeneration. We found a marked neurodegeneration in the amygdala and brainstem of 9-month-old SHRs, while CST and the GABAAR agonist Muscimol provided significant neuroprotection. Enhanced phosphorylation of Akt and ERK accounted for these neuroprotective effects through anti-inflammatory and anti-apoptotic activities. Overall our results point to CST exerting potent antihypertensive and neuroprotective effects plausibly via a GABAergic output, which constitute a novel therapeutic measure to correct defects in blood flow control in disorders such as stroke and Alzheimer's disease.

Brain excitatory/inhibitory circuits cross-talking with chromogranin A during hypertensive and hibernating states.

To date, many scientific attempts have been directed towards the development of experimental models for the identification of neuronal mechanisms evoking cardiovascular and hemodynamic dysfunctions. The spontaneously hypertensive rat (SHR), a genetic model of essential hypertension, has become a valuable rodent for the characterization of molecular markers in hypertensive-related diseases. Recently, growing interests have also been directed to a new experimental paradigm i.e. hibernation, a physiological state consenting the hamster (Mesocricetus auratus) to activate protective mechanisms against ischemic-like complications during the arousal phase. With this intention, the present review will focus attention on specific neurosignaling systems involved with the preservation of hemodynamic conditions in those brain areas that play a pivotal role on such a feature. It is widely known that healthy neurons conserve their structural and responsiveness properties in presence of a constant blood supply, which is assured by their coupling to microvessels and perivascular astrocytes as well as by secretory proteins such as chromogranin A (CgA). So, it will be interesting to establish if this protein alone or with the participation of excitatory/inhibitory neurosignals is capable of influencing some brain areas controlling cardiovascular conditions in both SHRs and hibernating hamsters. In this context, the present work will deliver the most important findings regarding neuronal CgA and its cross-talking ability with major inhibitory (GABA/adenosine) and/or excitatory (glutamate) neuroreceptor systems in relation to hypertensive/hypotensive states of both animal models. Indications deriving from such approaches may provide clinically useful insights regarding their role as protective factors of hemodynamic and neurological disorders.

Amygdalar excitatory/inhibitory circuits interacting with orexinergic neurons influence differentially feeding behaviors in hamsters.

Recently, environmental stimuli on different neurobiological events, via participation of distinct amygdalar (AMY) ORXergic fibers have aroused wide interests in view of their ability to modify neuronal linked stressful and physiological homeostatic conditions. Results of the present study indicate that ORXergic (ORX-A/B) circuits of the facultative hibernating golden hamster (Mesocricetus auratus) central AMY (CeA) and basolateral AMY (BlA) nuclei constitute major sites of feeding behaviors. Indeed, hamsters after treatment of BlA with ORX-A frequently ingested greater quantities of food as compared to controls, while ORX-B in CeA induced a very (p<0.001) great consumption of water. The same nuclei treated separately with either ORX-A or ORX-B ± the selective α(1) GABA(A) benzodiazepine receptor agonist (zolpidem) dedicated less time to eating and drinking sessions. Conversely, hamsters that received the same neuropeptides but this time with the glutamatergic agonist NMDA displayed greater hyperphagic effects above all for ORX-A. When behavioral changes were compared to the expression of the specific ORXergic receptor (ORX-2R), an up-/down-regulating pattern was detected in some limbic areas (AMY, hippocampus and hypothalamus) following treatment with ORX-A or ORX-B plus NMDA. Overall, indications deriving from this study strongly point to hamster BlA-enriched ORX-A fibers in combination with either inhibitory or excitatory signals as main targets of hyperphagic responses while CeA ORX-B activities in presence of these same neuronal signals predominantly induced drinking motivational behaviors. The distinct behavioral activities of these two neuropeptides may have useful clinical bearings toward psychiatric and sleeping disorders such as bulimia and narcolepsy.

Kidney transplantation at Annunziata Hospital of Cosenza: report of 10 years experience.

OBJECTIVE: Kidney transplantation represents the gold standard for treatment of patients with end-stage renal disease. Herein we sought to report our 10-year experience with cadaveric kidney transplantations. PATIENTS AND METHODS: From February 1995 to September 2008, we performed 115 kidney transplantations. Patients were followed for an average of 4.9 years (range, 2.2-10.6 years). The cold ischemia time (CIT) averaged 13 +/- 3 hours, while the mean warm ischemic time was 25 +/- 10 minutes. The ureteral-bladder anastomosis was performed using Bracci catheters in the first series of 72 transplants, and double-J stents in the other 41 cases. The average waiting time was 122 +/- 21 months. The immunological regimens were prescribed according to the American Society of Nephrology (K/DOQI) with reference to comorbidity and concomitant risk factors and reported drug toxicity events. We transplanted kidneys with anatomic variations, ie, multiple arteries and double veins, and one double transplant of marginal organs. RESULTS: Our overall complication rate was 9.18%. The 10-year patient and graft survival rates were 89% and 84%, respectively. The percentage of biopsy-proven acute rejection episodes was 22.16%, while chronic allograft nephropathy (CAN) accounted for 15.3% at 5 years. The incidence of delayed graft function (DGF) was 14.05%. Finally, we noted 3 cases of cardiovascular death. CONCLUSION: Our experience showed excellent patient outcomes compared with other Italian and European data.

The endocrine disruptor atrazine accounts for a dimorphic somatostatinergic neuronal expression pattern in mice.

It has now been established that a large number of man-made and natural chemicals are capable of interfering with the action of natural hormones. In this category "endocrine disruptors" such as the herbicide atrazine, when administered at ecological low doses (1 or 100 microg/kg per day) from gestational day 14 to postnatal day 21, provided a clear dimorphic neurodegenerative pattern in some brain areas of the domestic mouse (Mus musculus). Indeed, the high concentration (100 microg/kg per day) with respect to the low concentration (1 microg/kg per day) induced relevant neuronal damage in extrahypothalamic sites, such as the cortical and striatal areas in both sexes. Marked alterations in other areas, including the hippocampal and hypothalamic nuclei, were mostly typical of the female. At the neuronal level, the neuropeptide somatostatin, specific for the secretion of growth hormone, seemed to be a major target of atrazine effects, as demonstrated by evident subtype2,3,5 receptor mRNA differences of this neuropeptide, at least for the first two subtypes. In particular, a very strong (p < 0.001) upregulation of subtype2 expressing neurons was detected in female hypothalamic areas, specifically the suprachiasmatic nucleus, whereas a similar downregulatory trend was reported for some extrahypothalamic areas such as the striatum. Interestingly, very strong upregulatory and downregulatory actions were detected for neurons expressing subtype3 in male hypothalamic and amygdalar regions and in the cortical and hippocampal areas, respectively. Overall, it appears that these first neurotoxicological effects of atrazine are very likely linked to dimorphic expression patterns of specific somatostatin subtypes in discrete but key hypothalamic and extrahypothalamic areas of Mus musculus.

Morphological and functional variations of Leydig cells in testis of the domestic pig during the different biological stages of development.

The relationship of morphometrical and androgen receptor evaluations of the main testicular interstitium cellular element (Leydig cells) in the domestic pig provided interesting numerical and morphological features during the different aging stages. As early as 25 days (a period in which the pig is sexually immature) there was a low number of Leydig cells (1.46 x 10(8)) with respect to a 78% and 35% increase in the adult (2.48 x 108) and aged (1.78 x 10(8)) animal, respectively. Interestingly, when the volume density of Leydig cells was considered, the average volume of these cells seemed to be high (75%) in the aged pig with respect to the young immature animal whereas a lower increase (27%) was observed for the adult animal. Moreover, the evaluation of testosterone receptor binding sites in the testis at the various stages of development also displayed a differentiated pattern since elevated testosterone receptor binding levels of the high dissociation affinity type were obtained for the adult pig. Thus, from the combined morphological variations of Leydig cells and testosterone receptor binding activity, it appears that this androgenic receptor component exerts distinct autocrine effects on the different functional features of some testicular tissue constituents at the different aging stages of the domestic pig.

Dimorphic features of the different alpha-containing GABA-A receptor subtypes in the cortico-basal ganglia system of two distantly related mammals (hedgehog and rat)

This investigation represents a first study dealing with the dimorphic differences of the main alpha-containing gamma-aminobutyric acid (GABA(A)) receptors in the brain of two distantly related mammals (hedgehog and rat). The labeling of these receptors in the presence of zolpidem (highly selective benzodiazepine agonist) and under the different degree of GABA(A)/benzodiazepine allosteric coupling activities accounted for a heterogeneous colocalization of alpha1-enriched and of alpha2/3-enriched and alpha5-enriched GABA(A) receptors in some areas of the cortico-basal ganglia system (including the important ventrolateral thalamic station) of both mammalian sexes. In the hedgehog, the greatest (P<0.001) GABA-dependent reduction of zolpidem inhibition constants was mostly registered in alpha1- and/or alpha5-enriched areas, such as the frontoparietal cortex lamina III (235%), ventrolateral thalamic nucleus (128%), and substantia nigra pars reticulata (110%) of the male. However, the greatest reductions in the rat were instead detected in the male substantia nigra pars reticulata (192%) and female striatum (120%), areas which are enriched either by the colocalization of alpha1- with alpha2/3-subunits or by all three alpha-subunits. These results support the contention that a sex-related alpha-containing GABA(A) receptor sensitivity constitutes an important element in the execution of skilled motor activities during the different socio-sexual behaviors of the two mammals.

Dimorphic distribution of the two main GABA(A) binding sites in cortical and limbic areas of a rodent living in natural environmental conditions.

Labeling of the two more important gamma-aminobutyric acidA (GABA(A)) supramolecular sites with [3H] muscimol (GABA(A)) and [3H] flunitrazepam (benzodiazepine) provided saturable, stable, and dimorphic binding activities in cortical and limbic regions of the wood mouse Apodemus sylvaticus. Of the cortical layers, which contained the highest [3H] muscimol binding levels, only the female lamina V supplied a greater (51%; P <0.01) receptor density than in the male. Areas of the limbic system instead proved to be the more favorable targets for differential GABA(A) binding levels. The highest (P <0.001) and higher levels were found in the oriens-pyramidalis CA1 layer of the hippocampus (65%) and in the vertical limb diagonal band-medial septal nucleus (48%), basolateral amygdala nucleus (45%), and ventromedial hypothalamic nucleus (43%), respectively, of the female. A similar pattern was obtained for [3H]flunitrazepam binding activity, especially in the presence of GABA. The highest and higher binding activities were obtained in the female central amygdala nucleus (78%) and in the ventromedial hypothalamic nucleus (52%), basolateral amygdala nucleus (48%), and oriens-pyramidalis CA1 layer of the hippocampus (47%), respectively, whereas higher levels were observed only in the male vertical limb diagonal band-medial septal nucleus (56%). Even in the cortical regions, the female exhibited higher (42%; cortex lamina V) and moderately higher (38%; cortex lamina VI) levels, with binding differences in the latter site plus in the basolateral amygdala nucleus occurring in a GABA-nondependent manner. From the saturation binding analyses it was possible to reveal that both maximal number of binding sites (Bmax) and mean dissociation constant (K(D)) modifications were responsible for receptor differences of the two GABAergic sites. These findings tend to suggest that dimorphic variations of the GABA(A) supramolecular sites, in some cortical and limbic regions, are strongly involved in sex-specific aggressive and reproductive activities of rodents living in their natural habitats.

Effects of N omega-nitro-L-arginine methyl ester on benzodiazepine binding in some limbic areas of hyperlipidaemic rats.

Quantitative autoradiography techniques were used to evaluate the chronic effects of the potent nitric oxide synthase inhibitor, N omega-nitro-L-arginine methyl ester, on the binding pattern of [3H]flunitrazepam (benzodiazepine agonist) in some behaviorally key limbic areas of the genetic hyperlipidaemic Pittsburg Yoshida rat. Administration of this potent synthase inhibitor was capable of supplying higher and moderately higher binding levels in the basolateral amygdala nucleus (+52%) and in the oriens-pyramidalis CA1 hippocampus layer (+38%), respectively. When we tested for the binding changes in the presence of GABA (principal benzodiazepine modulator) we noticed that a physiological concentration (20 microM) of this inhibitory neurotransmitter was sufficient to induce notable changes in other limbic areas. In fact, lower binding values (-65%) were reported for the bed nucleus of stria terminalis whereas moderately higher values (+38%) were obtained for the radiatum-lacunosum molecular CA1 hippocampus layer. From the saturation studies, it was possible to observe that the major receptor variations provoked by the potent synthase inhibitor were not only due to changes in the total number of binding sites because there were variations, as in the case of the basolateral amygdala nucleus, that were instead due to differences in the affinity binding state. These results provide evidences of a GABAergic-nitric oxide synthase inhibitor interaction that might also be involved in the regulation of convulsive, anxiolytic, and aggressive behaviors that are modulated at the benzodiazepine site.

Sexual dimorphism of GABAA receptor levels in subcortical brain regions of a woodland rodent (Apodemus sylvaticus).

This is the first report of quantitative autoradiography results showing sex differences of GABAA receptor levels in brain regions of a wild rodent (wood mouse, Apodemus sylvaticus) living in its natural habitat. The labeling of this GABAergic site with its specific high affinity radioligand [3H] muscimol provided a heterogeneous and dimorphic binding pattern in some of the neural centers. In the female, higher (> or = 50 < or = 65%) to moderately higher (< 50%) binding levels than in the male, even after correction of the specific binding values using the calculated quenching coefficients, were observed in the substantia nigra pars reticulata and ventral lateral thalamic nucleus, brain centers that are relays of motor circuits. In the male, on the other hand, a higher level was only obtained in the caudateputamen. Relays of the stria terminalis-hypothalamic-central gray pathway such as the bed nucleus of the stria terminalis, the pontine central gray and the ventromedial hypothalamic nucleus, were among the other female brain areas with an extremely higher (> 65%) to higher and moderately higher binding activity than in the male. From the saturation analyses, it appeared that the binding differences were mainly due to Bmax variations, although closer examinations revealed that changes in the KD might have also accounted for [3H] muscimol binding differences, as shown by the high KD and Bmax values in the bed nucleus of the stria terminalis, the substantia nigra pars reticulata and the pontine central gray of the female wood mouse. These findings suggest that the dimorphic binding activity of GABAA receptors in the above brain regions might be involved in neuronal circuitry mechanisms related to sex-specific social behaviors in rodents living in their natural environmental conditions.

Comparison of melatonin-binding sites in the brain of two amphibians: an autoradiographic study.

Neuroanatomic comparison of the binding capability of 2-[125I] iodomelatonin in the crested newt Triturus carnifex Laur. and the green frog Rana esculenta, using quantitative autoradiographic techniques, revealed a heterogeneous distribution pattern. The highest and relatively high binding activities were shown to occur in the optic tracts and in the suprachiasmatic area of the hypothalamus and the optic tectum, respectively, of both species. Low or no 2-[125I] iodomelatonin binding values were obtained in the preoptic nucleus, the tuberal hypothalamus, the medulla oblongata, the septum and the dorsal pallium. A differential binding pattern was observed in the amygdaloid nucleus pars lateralis, the striatum and the hindbrain of these amphibians. Indeed, notably high binding levels were shown to occur in the former two brain areas of the crested newt, whereas high levels were displayed in the latter brain region of the green frog. On the basis of elevated quantities of melatonin receptors in mesencephalic, hypothalamic and telencephalic sites, it seems plausible to ascribe some important sensory functions to this receptor system in both species. The remarkably different binding activities in the brain of the two amphibians could be correlated with the simpler cytoarchitectonic brain structure of urodeles and with species-specific variations.

Combined gonadal and photic influences on 2-[125I] iodomelatonin-binding level changes in some brain areas of the quail.

Quantitative receptor autoradiography was used to study the binding of 2-[125I] iodomelatonin in the brain of the castrated and gonadally intact male Japanese quail Coturnix japonica exposed to both long- and short-day photoperiod cycles. The distribution study displayed that these conditions were responsible for a heterogeneous binding pattern as shown by elevated receptor levels being located in visual brain centers, such as the stratum opticum, nucleus pretectalis, and nucleus geniculatus lateralis, pars ventralis, while lower values were found in the nucleus lateralis hypothalami and nucleus isthmi pars magnocellularis. Closer examination of the 2-[125I] iodomelatonin-binding pattern following the different gonadal and photic influences showed that combination of the gonadally intact condition and a 16L:8D (long-day) photoperiod cycle was required for the greater binding changes. These differences occurred in brain sites such as the area preoptica, paleostriatum primitivum, and nucleus ectomamillaris. Saturation binding studies, which were carried out only in some of the above areas, revealed that the combined gonadal- and photic-induced changes are basically due to the modifications of total number of binding sites. The importance of a gonadal steroid modulatory role in the photic-dependent melatonin binding activity suggests that other types of neuronal mechanisms might be involved in the regulation of neuroendocrine and socio-sexual behaviors in nonmammalian vertebrates.

Gonadal-GABAergic interaction is an important factor involved in photoperiod-induced 2-[125I] iodomelatonin binding changes in the Japanese quail brain.

The type of mechanism(s) by which melatonin alone and/or through the intervention of other putative neurotransmitters is able to control circadian rhythms remains unresolved. Comparison of 2-[125I]iodomelatonin binding pattern in the brain of castrated and gonadally intact Japanese quail (Coturnix japonica), using quantitative receptor autoradiography, displayed that the combination of the intact gonadal condition and a long-day (16L:8D) photostimulatory schedule is responsible for major binding changes. In fact, high and low binding levels were obtained in the suprachiasmatic area and nucleus ectomamillaris (p < 0.01) and in the nucleus preopticus anterior and paleostriatum primitivum (p < 0.001), respectively. A gonadal modulatory role was not always evident in all brain areas as revealed by long-day photic cycles producing diminished (p < 0.01) binding levels in the anterior neostriatum and the nucleus rotundus of both castrated and gonadally intact animals, although elevated values were also found in the substantia grisea centralis (p < 0.05) of the same animals. Saturation binding studies revealed that gonadal and/or photic effects induce alterations in the number of binding sites, whereas the affinity constant varied only in some hypothalamic sites. Testing of GABAergic activity on 2-[125I] iodomelatonin binding levels showed that this inhibitory neurotransmitter was responsible for increasing low receptor values. Moreover, GABA-dependent influences were shown to be mediated via a GABAA receptor subtype since bicuculline (specific antagonist of this site) inhibited the elevated GABA-induced melatonin binding levels in the above brain sites of the gonadally intact quail exposed to both photoperiod cycles. Even in this case, melatonin binding changes were due to the variations in the number of binding sites. The apparent GABAergic-gonadal influence resulting in changes of the 2-[125I] iodomelatonin binding values, under the different photic conditions, provides evidences of other probable neural mechanisms that entrain circadian rhythmicity in neuroendocrine activities and in sociosexual behaviors.

ANF binding sites in the heart of the quail (Coturnix coturnix japonica).

The distribution pattern of rat [125I]-atrial natriuretic factor (ANF) binding sites in the cardiac regions of the Japanese quail was examined by in vitro quantitative autoradiography. Elevated ANF binding densities (519 +/- 121 fmol/mg protein) were found in the posterior vena cava, while lower binding levels (between 40 and 50 fmol/mg protein) were found in sinus venosus, aortic bulb, and endomural vessels, with the ventricular wall having the lowest value (17.6 +/- 8.8 fmol/mg protein). Scatchard analyses of the ANF binding characteristics (Kd, Bmax) revealed both low (94 +/- 55 fmol/mg protein) and high (1161 +/- 69 fmol/mg protein) Bmax values. Receptors with higher Kd values than those observed in other cardiac regions (Kd between 30 and 60 pM) were found in the vena cava and in the heart ventricle (Kd between 113.2 and 229 pM).

Steroid hormones and receptors of the GABAA supramolecular complex. I. Benzodiazepine receptor level changes in some extrahypothalamic brain areas of the female rat following sex steroid treatment.

The effects of sex steroid hormones on the different receptor binding sites of the GABAA molecule remain unclear. In this report we have demonstrated, using autoradiography techniques, that the distribution pattern of the benzodiazepine receptors (a component of the GABAA molecule) in some extrahypothalamic brain regions is altered by both in vivo and in vitro sex steroid hormone treatment. In vivo administration of the sex steroids estradiol and progesterone induced a significant change in [3H]flunitrazepam (benzodiazepine agonist) binding levels in the amygdala, and cortico and posterior brain nuclei of the female rat. In fact, elevated and diminished receptor-binding levels were obtained in the corticomedial amygdala nucleus and in the pontine central gray matter respectively, following the administration of estradiol. Significant hormonal effects were also shown for animals that received only a progesterone dose, as demonstrated by the increased and decreased receptor levels in the basolateral amygdala nucleus and cortex lamina VI and in the substantia nigra pars reticulata, respectively. It was interesting, at this point, to investigate whether the hormone effects on [3H]flunitrazepam binding changes might be mediated through a GABA-dependent activity, because the benzodiazepine and GABAA receptors are coupled to a chloride ion channel in an allosteric manner. When 50 microM GABA was added to the incubation medium, substantially altered binding levels were recorded in animals that received progesterone replacement therapy only. The GABA-induced progesterone effects both increased substantially the binding levels in the oriens-pyramidalis CA1 layer of the hippocampus and in the intermediate gray layer of the superior colliculus as well as reducing receptor levels in the substantia nigra pars reticulata.(ABSTRACT TRUNCATED AT 250 WORDS)

Steroid hormones and receptors of the GABAA supramolecular complex. II. Progesterone and estrogen inhibitory effects on the chloride ion channel receptor in different forebrain areas of the female rat.

The inhibitory effect of female sex steroid hormones on the binding of [35S]t-butylbicyclophosphorothionate [35S]TBPS to the chloride ion channel receptor in different forebrain areas of the female rat proved to be of a differential nature. The in vivo administration of estradiol and estradiol+progesterone were responsible for substantially lower chloride ion channel receptor levels in brain areas that contain elevated steroid receptors, such as the medial preoptic area, the cortico-medial amygdala nucleus, the vertical limb diagonal band-medial septal nucleus and the cortex lamina V. The administration of progesterone alone reduced receptor levels in the oriens-pyramidalis CA1 layer of the hippocampus, caudate putamen, cortex lamina VI (brain areas that contain little if any steroid receptors) and in the lateral and basolateral amygdala nucleus (brain sites that contain noninducible progesterone receptors). On the basis of the progesterone-inhibitory activity on the chloride ion channel receptors, it was important to investigate whether progesterone per se or whether the potent progesterone metabolites 3 alpha-hydroxy-5 alpha-dihydroprogesterone (3 alpha, 5 alpha-THP) and 3 beta-hydroxy-5 beta-dihydroprogesterone (3 beta,5 beta-THP) were involved in the binding level changes, and to establish the specific brain sites where these steroid effects occur. In fact, in vitro addition of the 5 alpha-reduced progesterone metabolite produced even greater depressive effects on [35S]TBPS binding not only in the same brain areas as the in vivo progesterone replacement therapy but also in some sites that provided significant receptor level changes following the sequential administration of estradiol+progesterone. However, when the 5 beta-reduced metabolite was tested on the binding of [35S]TBPS to the chloride ion channel receptor, only the basolateral amygdala nucleus, the cortex lamina VI and the dorsolateral septal nucleus exhibited changes. Because the steroid-mediated chloride ion flux is regulated in a GABA-dependent manner, we next checked for the type of GABA effects on the chloride ion channel receptor levels and found that GABA not only intensified the 3 alpha,5 alpha-THP inhibitory effects but, together with this progesterone metabolite, was also involved in binding changes in the vertical limb diagonal band-medial septal nucleus. It is interesting to note that the GABA effects on 5 beta-metabolite-induced receptor changes were not of the enhancing type, but tended, rather, to be inhibitory.(ABSTRACT TRUNCATED AT 400 WORDS)

The neuroanatomic binding pattern of [125I]atrial natriuretic factor in the Japanese quail brain.

Application of quantitative autoradiography technique provided a discrete anatomical distribution pattern of the atrial natriuretic factor (ANF) in the Japanese quail, Coturnix coturnix japonica, brain. The highest binding levels of [125I]ANF were shown to occur in telencephalon areas, such as fasciculus diagonalis Brocae (232 fmol/mg protein), septum (194 fmol/mg protein) and olfactory bulb (153 fmol/mg protein), and in posterior sites, such as nucleus interpeduncularis (177 fmol/mg protein), while lower levels (> 51 < 87 fmol/mg protein) were found in the hypothalamic sites of the diencephalon. The similar ANF receptor density levels in some brain areas of the quail as well as both mammalian and non-mammalian species suggest that this peptide might be involved in osmoregulatory activities (at the brain level) and furthermore indicate a probable functional conservation of ANF in vertebrates.

A quantitative autoradiographic study of GABAA and benzodiazepine receptors in the brain of the frog, Rana esculenta.

Specific binding sites for GABAA and benzodiazepines were detected in the brain of the frog Rana esculenta after the in vitro incubation of tissue sections with their respective specific agonists [3H] muscimol and [3H] flunitrazepam. Conditions for the binding assay were optimized and as a result binding was saturable and specific. Quantitative autoradiographic receptor measurements in the different brain sections showed that elevated levels of [3H] muscimol binding sites were found in the two layers of the cerebellum (periventricular and external) with the highest binding densities being detected in the periventricular layer. Relatively high densities of [3H] muscimol binding sites were also observed in the torus semicircularis of the mesencephalon and in the thalamic nucleus rotundus and posterolateral nucleus, plus the mitral cell layer of the olfactory bulb, the amygdala pars lateralis and the striatum of the telencephalon. Intermediate to low binding levels were obtained in the remaining brain areas such as the external layer of the optic tectum, the dorsomedial and dorsolateral anterior thalamic nuclei, the medial and lateral pallium, the medial septal nucleus, the preoptic nucleus, the dorsal and ventral infundibular nuclei of the hypothalamus and the interpeduncular nucleus. Autoradiographic evaluation of benzodiazepine receptors revealed that binding levels of [3H] flunitrazepam were overall lower than those of the GABAA sites. In fact the highest [3H] flunitrazepam binding levels were observed in the striatum, external layer of the optic tectum and the torus semicircularis.(ABSTRACT TRUNCATED AT 250 WORDS)

A quantitative autoradiographic study of 125I atrial natriuretic factor in the heart of a teleost fish (Conger conger).

Quantitative receptor autoradiographic study of 125I-atrial natriuretic peptide factor (ANF) in the heart of a teleost fish Conger conger has shown that a heterogenous distribution of 125I-ANF binding exists in the different cardiac regions. Elevated ANF binding densities (3,790 fmol/mg protein) were encountered in the innermost layer (tunica intima) of the bulbus arteriosus while lower binding levels (293-403 fmol/mg protein) were revealed in atrium and ventricle. In order to determine 125I-ANF binding characteristics (KD, Bmax) in the above cardiac sites, saturation binding assays were carried out. The results show that low 125I-ANF KD values (28.8-52.6 pM) were found in the atrium and in the bulbus arteriosus with respect to the higher KD values (373 pM) of the ventricle. The number of binding sites were respectively 632 and 1,279 fmol/mg protein for the atrium and the ventricle, while a substantially elevated Bmax of 7,235 fmol/mg protein was found for the bulbus arteriosus. These results may furnish some insights concerning ANF receptor binding activity and its putative regulatory role of different cardiac functions.