Localization of 125I-insulin binding sites in the rat hypothalamus by quantitative autoradiography

In vitro autoradiography and computer video densitometry were used to localize and quantify binding of 125I-insulin in the hypothalamus of the rat brain. Highest specific binding was found in the arcuate, dorsomedial, suprachiasmatic, paraventricular and periventricular regions. Significantly lower binding was present in the ventromedial nucleus and median eminence. The results are consistent with the hypothesis that insulin modulates the neural regulation of feeding by acting at sites in the hypothalamus.     

Apolipoprotein E uptake and degradation via chloroquine-sensitive pathway in cultivated monkey cells overexpressing low density lipoprotein receptor

Alzheimer's disease with late onset is associated with inheritance of isoform 4 of the polymorphic apolipoprotein E (ApoE). We describe an experimental model where intracellular interaction between ApoE isoforms and cellular proteins may be studied. Internalization and intracellular fate of ApoE from fresh normal cerebrospinal fluid is followed in COS cells (an SV40 transformed Simian kidney cell line) overexpressing the low density lipoprotein receptor. Chase and chloroquine experiments strongly suggest that internalized ApoE travels through the endosomal-lysosomal pathway after dissociation from the receptor in early endosome.     

Heart rate and oxygen uptake response to angiotensin in the squirrel monkey at 10 degrees C.

Unanesthetized squirrel monkeys exposed to an ambient temperature of 10 degrees C showed elevations in total body oxygen consumption (VO2), systemic arterial blood pressure (BP), and heart rate (hr) above values recorded at 28 degrees C. Further elevation of BP in the cold by intravenous infusion of phenylephrine (5-50 microgram/kg-min) was accompanied by reduction in both VO2 and HR, and the changes in VO2 were proportional to those in HR. When BP was raised by intravenous infusion of angiotensin (0.05-1.0 microgram/kg-min), large elevations in BP were again accompanied by reductions in HR and VO2. However, for equivalent elevations in BP, the depressions in both HR and VO2 were much smaller with angiotensin than they were with phenylephrine. Previous studies in this laboratory have demonstrated that in response to experimental elevation of BP, reflexes originating at the sinoaortic baroreceptors depress not only HR but also VO2. The present results suggest that angiotensin modulates baro-reflexive responses to elevation in BP. The reductions in HR and VO2 that ordinarily occur in response to baroreceptor stimulation may be modified by an action of angiotensin on the central nervous system.     

Olfactory discrimination of squirrel monkey mothers by their infants.

Infant squirrel monkeys, 8 to 24 weeks of age, were tested for their ability to discriminate their own mother from another lactating mother by means of olfactory cues, visual cues, or a combination of both. Discrimination by olfactory cues was unequivocal. Addition of static visual cues did not enhance discrimination, and infants showed little evidence of discriminating the mother by means of static visual cues alone. These results suggest that olfaction may be a more significant factor in primate early attachment than has generally been assumed.     

Coxsackievirus B4 nephritis in the squirrel monkey.

Seventeen squirrel monkeys (Saimiri sciureus) were experimentally infected with Coxsackievirus B4, and the kidneys, as well as other organs, were studied for pathological changes induced by the virus. Seven (41%) of these monkeys developed renal lesions--interstitial and glomerular. The Coxsackievirus was identified in 4 of these 7 monkeys (by isolation from the renal tissue in 2, by immunofluorescence staining of viral antigen in 1, and by electron microscopic finding of viral particles in 1). The renal lesions produced by Coxsackieviral infection described in this report resemble those seen in renal disease in man. These findings support the concept that viruses can produce glomerular and interstitial renal disease. This report also describes a good animal model for the study of viral disease of the kidney.     

Experimental Lassa virus infection in the squirrel monkey.

Experimental Lassa virus infection was investigated in a nonhuman primate in order to elucidate the target organs of the viral infection and the course of pathologic events. Four squirrel monkeys (Saimiri scirreus) were inoculated intramuscularly with Lassa virus and sacrificed for organ titrations and histopathology, one each day, on Days 7, 12, 14, and 28 after inoculation. The animals showed a variable clinical course, with an incubation period of 8 to 18 days. The virus was demonstrated to be virtually pantropic; however, lymph node, liver, and kidney were key early targets. After the onset of overt disease, patterns of lymphoreticulotropism, hepatotropism, nephrotropism, adrenotropism, and persistent viremia were evident. Complement-fixing antibody failed to develop after 28 days of infection. Histopathologic findings included germinal center necrosis in spleen and lymph node; myocarditis; acute arteritis; renal tubular necrosis and regeneration; hepatocytic regeneration; chronic inflammation of choroid plexus, ependyma, and meninges; and cerebral perivascular cuffing. There is a relationship between many of these lesions and certain features of other arenavirus infections. The model offers the opportunity to pursue investigations of experimental pathogenesis, transmissibility, and efficacy of immunotherapy.     

Motion sickness-induced food aversions in the squirrel monkey.

Conditioned aversions to colored, flavored water were established in Squirrel monkeys (Saimiri sciureus) by following consumption with 90 min of simultaneous rotational and vertical stimulation. The experimental group (N = 13) drank significantly less of the green, almond-flavored test solution than did the control group (N = 14) during three post-treatment preference testing days. Individual differences were noted in that two experimental monkeys readily drank the test solution after rotational stimulation. Only two of the experimental monkeys showed emesis during rotation, yet 10 monkeys in this group developed an aversion. These results that (1) motion sickness can be readily induced in Squirrel monkeys with simultaneous rotational and vertical stimulation and (2) that conditioned food aversions are achieved in the absence of emesis in this species.     

Ontogeny and characterization of [125I]Bolton Hunter-eledoisin binding sites in rat spinal cord by quantitative autoradiography.

The distribution and characteristics of [125I]Bolton Hunter-eledoisin binding sites in rat lumbar spinal cord were studied during postnatal development by in vitro receptor autoradiography. At three, six and 10 days of age, specific [125I]eledoisin binding was distributed throughout the dorsal and ventral horns of the spinal cord. In contrast, from day 24 onwards, specific binding of [125I]eledoisin was confined to superficial layers of the dorsal horn, with negligible amounts of specific binding in the ventral horn. [125I]Eledoisin binding to neonatal (three day) and adult (eight to 12 weeks) spinal cord sections was characterized using tachykinin agonists. In both dorsal and ventral horns of neonatal spinal cord, the rank order of potency of agonists indicated that the majority (64%) of specific [125I]eledoisin binding was to neurokinin-3 binding sites. The identity of the non-neurokinin-3 sites labelled by [125I]eledoisin remains to be determined. In adult rat spinal cord, [125I]eledoisin appeared to bind exclusively to neurokinin-3 binding sites. These results suggest that major changes take place in the localization of neurokinin-3 receptors during postnatal ontogeny of the rat spinal cord. These changes may reflect an important role for tachykinins in neuronal plasticity of the developing spinal cord.     

Decreased uptake of low density lipoprotein by LLC-PK cells cultured at low magnesium concentration.

LLC-PK cells grown in tissue culture on medium containing normal or deficient amounts of magnesium were presented with 125I-low density lipoprotein (LDL) and the uptake determined. Total LDL uptake by Mg-deficient cells decreased as the magnesium concentration decreased. The receptor binding of LDL and the internalization of LDL by the Mg-deficient cells were reduced. Degradation of LDL by Mg-deficient cells, when corrected for the reduced uptake, was less affected.     

Diagnosis of pregnancy in the squirrel monkey

Diagnostic pregnancy assessments have been carried out in 18 squirrel monkey females using Delfs' method for chorionic gonadotropin bioassay palpatory, x-ray and other auxiliary techniques. Out of the 24 bioassay tests performed, 11 were positive (one of them weakly positive) and 13 negative. These results were confirmed by the other parallel measures, except in two cases of possible false positive reactions. No false negative results were noted. The implications of the results are discussed in terms of the sensitivity and reliability of the bioassay in the early diagnosis of pregnancy in the squirrel monkey.     

Measurement of squirrel monkey serum IgG levels by a two-site sandwich radioimmunoassay with monoclonal antibodies

Monoclonal antibodies against the squirrel monkey Saimiri sciureus IgG have been produced for a more specific analysis of the antibody-related immunological aspects in experimental human or monkey malaria. Two monoclonal antibodies, 3D8/D5 and 3F11/G10, out of 64 reacted with distinct epitopes on the IgG present throughout the complete population without interfering with each other. The 2 monoclonal antibodies were used to develop a highly specific, reliable and sensitive two-site sandwich radioimmunoassay for the measurement of the serum IgG levels in 83 animals. The antibodies also allowed us to produce by a simple immunoabsorbent technique a highly purified IgG standard easy to calibrate and store. The assay permits the detection of IgG levels as low as 0.48 ng/ml. The standard curve is linear between 3.9 and 125 ng protein/ml and allows by a simple mathematical equation an accurate measurement of the serum IgG levels.     

Pharmacological analysis of 125I-Bolton and Hunter labelled eledoisin binding sites in rat spinal cord by quantitative autoradiography

Using the technique of quantitative autoradiography it has been possible to investigate and compare the pharmacological characteristics of 125I-Bolton-Hunter conjugated eledoisin (125I-BHE) binding sites in rat spinal cord with those in the rat cortex. 125I-BHE specific binding sites were discretely localised in the outer layers of the rat spinal cord. The rank order of affinity of unlabelled tachykinins in competing for 125I-BHE specific binding sites in rat spinal cord (NKB greater than ELE greater than NKA greater than SP) was identical to that found in the rat cortex suggesting the presence of 'NK-3 like' receptors in rat dorsal horn.     

Analysis of fluorescent low density lipoprotein uptake by lymphocytes, paradoxical increase in the elderly

Uptake of dioctadecylindocarbocyanine (DiI)-labelled low density lipoproteins (LDL) by peripheral blood lymphocytes (PBL) from young healthy donors has been characterized by flow cytometric analysis. The receptor positive cells were primarily (> 70%) T cells. Saturation and competition studies were performed with freshly isolated PBL as well as after a 2–3-day incubation in cholesterol-free medium. In both cases uptake was specific for LDL and not high density lipoprotein. It was also abrogated by chemical modification of apo B, and was not shown by PBL from a patient with familial hypercholesterolemia.

DiI-LDL-uptake by cells from elderly donors was compared with that of PBL from young donors. There was a clear increase in uptake by freshly isolated PBL from aged donors which was shown not to stem from underlying “sickness”. In contrast, uptake by pre-incubated cells was very variable, in terms of percentage receptor-positive cells and the level of uptake by those cells. However, LDL rescued mevinolin-suppressed mitogen responses from both old and young donors indicating that there is no impairment of uptake or degradation of LDL by PBL from the elderly.     

Social status constrains the stress response in the squirrel monkey.

The influence of dominance on the pituitary-adrenal and gonadal systems was evaluated in male squirrel monkeys. Basal and stress levels of plasma cortisol and testosterone were determined in eight male pairs across a 5-week period. The data indicated that squirrel monkeys have unusually high levels of steroid hormones in comparison to other species. Dominant males had higher levels of cortisol and testosterone and showed a smaller stress response than did subordinate males.     

High density lipoprotein uptake by freshly isolated human peripheral blood T lymphocytes

A high density lipoprotein (HDL) receptor/binding site has been identified on peripheral blood lymphocytes, and some of its properties were compared with those of HDL receptors on other cell types. Binding studies were performed using fluorescent (dioctadecylindocarbocyanine)-labelled HDL (DiI-HDL) and analyzed by fluorescence microscopy and quantitative flow cytometry. Uptake of low levels of DiI-HDL during a 2 h incubation at 37 degrees C was a property of all lymphocytes, i.e. not of one particular subset only. Visual inspection of these cells in the fluorescence microscope revealed both membrane and cytoplasmic fluorescence, indicating that DiI-HDL become internalized during the 2 h incubation; internalization appeared to be a receptor-mediated process. In competitive binding studies, apo E-free HDL competed effectively for DiI-HDL binding, whereas LDL competed very weakly. Two features of DiI-HDL uptake are demonstrated which are unique to lymphocytes: 1) it was enhanced 3-6-fold by inclusion of EDTA in the incubation medium or by incubating in Ca2+/Mg2+ free medium, and 2) it was saturable at 37 degrees C.     

Cerebral representation of vocalization in the squirrel monkey

Summary Specific vocalization types following electrical stimulation of 5940 electrode positions are studied in 39 squirrel monkeys. Except cerebellum, caudal medulla, and a few cortical areas, the sites of stimulation were distributed throughout the brain. Each vocalization elicited was tested for reproducibility at the site of stimulation and in homologue structures. All vocalizations were analyzed spectrographically and then classified for eight fundamental vocalization types.The cerebral distribution of two call types forms continuous systems extending from midbrain via diencephalon into forebrain; the remaining call types are represented in several separate areas not continuous with each other. In medulla and pons the responsive substrates for vocalization follow the course of the spinothalamic tract; in midbrain they lie within the periaqueductal gray, lateral tegmentum, and lemniscus medialis; in diencephalon they are found in the hypothalamus and midline thalamus; in forebrain, finally they are distributed over amygdala, stria terminalis, substantia innominata, preoptic region, septum, rostral hippocampus, posteromedial orbital cortex, cingulate gyrus, and rostroventral temporal cortex. Hence a close relation between limbic system and vocalization producing substrates emerges.Among the brain structures yielding vocalization the mesencephalic periaqueductal gray is assumed to be the region where the electrical stimulus interferes most directly with specific vocalization mechanisms.Besides the anatomical site of stimulation the set of stimulus parameters is important for the elicitation of vocalizations. Relations between amplitude, frequency, and duration of impulses on the one hand and type, loudness, rhythm, duration, and latency of vocalization on the other hand were tested. The influence of the stimulus set on the reaction parameters depends also on the relative position of the electrode within the effective structure. Proper manipulation of stimulus parameters often results in the disintegration of a complex stimulus response into single components.Abbreviations a Nucl. accumbens - aa Area anterior amygdalae - ab Nucl. basalis amygdalae - ac Nucl. centralis amygdalae - al Nucl. lateralis amygdalae - am Nucl. medialis amygdalae - an Nucl. anterior thalami - anl Ansa lenticularis - aq Substantia grisea centralis - bc Brachium conjunctivum - ca Caudatum - cc Corpus callosum - cen Nucl. centralis superior (Bechterew) - cent Centrum medianum - ci Capsula interna - cin Cingulum - cl Claustrum - coa Commissura anterior - coli Colliculus inferior - cols Colliculus superior - cr Corpus restiforme - csp Tr. corticospinalis - db Fasc. diagonalis Brocae - dbc Decussatio brachii conjunctivi - f Fornix - gc Gyrus cinguli - gl Corpus geniculatum laterale - gm Corpus geniculatum mediale - gr Gyrus rectus - gs Gyrus subcallosus - h Campus Foreli - ha Nucl. habenularis - hi Tr. habenulointerpeduncularis - hip Hippocampus - hya Area hypothalamica anterior - hyp Area hypothalamica posterior - hyv Area hypothalamica ventralis - in Nucl. interpeduncularis - lap Nucl. lateralis posterior thalami - lav Nucl. lateralis ventralis thalami - le Lemniscus lateralis - lem Lemniscus medialis - lm Fasc. longitudinalis medialis - m Corpus mamillare - md Nucl. medialis dorsalis thalami - mt Tr mamillothalamicus - nst Nucl. striae terminalis - oi Nucl. olivaris inferior - ol Fasc. olfactorius (Zuckerkandl) - os Nucl. olivaris superior - p Pedunculus cerebri - pmc Brachium pontis - po Griseum pontis - pp Nucl. praepositus hypoglossi - pro Area praeoptica - pu Nucl. pulvinaris thalami - put Putamen - re Formatio reticularis tegmenti - rep Nucl. reticularis tegmenti pontis - rl Nucl. reticularis lateralis myelencephali - rub Nucl. ruber - s Septum - sm Stria medullaris - sn Substantia nigra - st Stria terminalis - sto Stria olfactoria lateralis - tec Tr. tegmentalis centralis - trz Corpus trapezoides - va Nucl. ventralis anterior - vpl Nucl. ventralis posterolaterali thalami - vpm Nucl. ventralis posteromedialis thalami - zi Zona incerta - II Tr. options - IICh Chiasma nervorum opticorum - III N. oculomotorius Nucl. nervi oculomotorii - IV N. and Nucl. nervi trochlearis - VI N. abducens - VII N. facialis - VIII N. acusticus - IX N. glossopharyngeus     

Glutamate carboxypeptidase II levels in rodent brain using [125I]DCIT quantitative autoradiography

The ability to visualize quantitatively glutamate carboxypeptidase II (GCPII) levels in vivo could advance our understanding of its function in health and disease. In the current study, we synthesized and evaluated a radiolabeled (iodine-125) analog of N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl]-S-3-iodo-L-tyrosine (DCIT), a potent antagonist of GCPII activity. We examined the regional distribution of [125I]DCIT binding in the rodent brain using quantitative autoradiography in order to confirm the validity of this radioligand as a marker of GCPII in the brain. The ultimate goal is to develop an imaging agent for assessing GCPII levels in the living brain. The specific binding of [125I]DCIT to rat brain followed a regional distribution consistent with previous studies describing regional brain GCPII gene expression and activity. We found a modest rostrocaudal gradient in which specific binding of [125I]DCIT to GCPII was lowest in cortical regions, with increasing levels of binding in midbrain structures and high levels of binding in hindbrain and brainstem. Autoradiography of [125I]DCIT in GCPII knockout and wild type mouse brain showed a gene-dose dependency confirming the selectivity of this radioligand for GCPII. We propose that [125I]DCIT is a selective radioligand that can be used to quantify brain GCPII levels in vitro using quantitative autoradiography.     

Physiologic responses to intravenous serotonin in the awake squirrel monkey.

Intravenous injections of serotonin in the monkey produces bradycardia and arrythmias, apnea followed by tachypnea, urination, and penile erection. These effects were not altered by anesthesia. Special consideration is given to the role of chemical and neural factors in the genital response, a reaction heretofore not noted in other species. An assortment of evidence suggests that this response is attributable to the local effect of serotonin on the genital vasculature.