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D W Pfaff

Publications and source records attributed to D W Pfaff.

At least 307 records · Page 17Linked to original sources
PubMed1986

Regional specificity in estradiol effects on [3H]uridine incorporation in rat brain.

In this study, the effects of 15 days of estradiol (E2) on tritiated-uridine incorporation were autoradiographically examined, on a cell-by-cell basis, in 4 E2-concentrating regions of the female rat brain. These areas included the ventromedial hypothalamus and medial amygdala nucleus, 2 regions involved in the behavioral components of reproduction, and the medial preoptic area and arcuate nucleus of the hypothalamus, 2 regions involved in the endocrine components of reproduction. Chronic E2 caused a 50% and 52% increase in tritiated-uridine incorporation in the arcuate nucleus and medial preoptic area, but was without effect in the ventromedial hypothalamus and medial amygdala nucleus. Somal area was also increased with E2 in the arcuate nucleus and medial preoptic area (16% and 43%, respectively) but not in the ventromedial hypothalamus and medial amygdala nucleus. The results indicate that the effects of estradiol on levels of newly synthesized RNA vary in a functionally and regionally specific manner within the brain.

Amygdala↗
PubMed1986

Cellular localization of proenkephalin mRNA in rat brain: gene expression in the caudate-putamen and cerebellar cortex.

The cellular locations of proenkephalin mRNA have been determined for the caudate-putamen and cerebellar cortex of the rat brain by in situ hybridization. In the caudate-putamen, more than half of the neurons express the proenkephalin gene. Morphologically, they are medium-sized cells that may represent projection neurons. In the cerebellar cortex, proenkephalin mRNA is present in a subpopulation of neurons in the granule layer that appear to be Golgi cells--i.e., inhibitory interneurons. The presence of [Met]enkephalin, a pentapeptide derived from proenkephalin, in these two brain areas is consistent with a synthetic role for this mRNA and implicates proenkephalin gene expression in the control of motor function.

Animals↗
PubMed1986

Single-unit activity of hypothalamic arcuate neurons in brain tissue slices. Effects of anterior pituitary hormones, cholecystokinin-octapeptide, and neurotransmitters.

Extracellular single-unit activity was recorded from hypothalamic arcuate nucleus (ARC) in brain tissue slices. Adult female Sprague-Dawley rats, ovariectomized or ovariectomized plus estrogen treated for at least 1 week, were used. Resting activity and responses of ARC neurons to six anterior pituitary hormones, or (as a positive control) cholecystokinin-octapeptide sulfate (CCK-8S), and a battery of four neurotransmitters including norepinephrine, serotonin, dopamine, and glutamate were recorded. A total of 263 neurons were recorded. Estrogen treatment did not cause any significant changes in the firing patterns nor in responses to most agents tested except for CCK-8S. A large percentage of the ARC neurons were either silent (40%) or slow-firing units (43% fired less than twice/s). Only a small percentage of ARC neurons (20-30%) responded to the anterior pituitary hormones, and these responses were small, delayed increases in firing despite the fact that CCK-8S stimulated more than half of the neurons with large responses. Glutamate was also excitatory, but not quite as effective as CCK-8S. Norepinephrine and serotonin were equally effective in eliciting a neuronal response (over 70% of units responded with an excitation or inhibition). Dopamine acted like norepinephrine, but was less potent. Since anterior pituitary hormones only weakly affected ARC neurons, these electrophysiological data give scant support to the notion that short-loop feedback is accompanied by electrical changes. In the ARC, however, CCK-8S may play some functional roles that are influenced by estrogen.

Adrenocorticotropic Hormone↗
PubMed1986

Brattleboro rat hypothalamic neurons transcribe vasopressin gene: evidence from in situ hybridization.

In situ hybridization has been used to identify specific hypothalamic magnocellular neurons, in normal and Brattleboro rats, that contain vasopressin (VP) or oxytocin (OT) mRNA. The subnuclear distribution of identified neurons in hypothalamic magnocellular nuclei after hybridization with several probes specific for OT or VP mRNA was in direct agreement with immunocytochemical descriptions of the distribution of cells containing VP or OT hormone. The number of grains per cell suggested that Brattleboro rats contained greater levels of OT mRNA, while hybridization with VP probes produced fewer grains in tissue from Brattleboro rats compared to normal rats. This paper provides the first cell-by-cell description of VP gene expression in the Brattleboro rat and demonstrates that VP gene transcription is confined precisely to the magnocellular neurons that normally synthesize VP hormone in normal rats.

Animals↗
PubMed1986

Combination of immunocytochemistry and in situ hybridization in the same tissue section of rat pituitary.

A procedure is described for combining avidin-biotinylated horseradish peroxidase immunocytochemistry, for localizing peptides or proteins, with in situ hybridization for localizing mRNA autoradiographically in the same cryostat section of paraformaldehyde-fixed rat pituitary. Protection against enzymatic degradation of target mRNAs during the immunocytochemical step was necessary and was accomplished by including an RNase inhibitor, 0.04% diethylpyrocarbonate, in primary and secondary antisera. This combination of methods may be useful in other tissues, as well, for (a) determining the relation of protein content to the concentration of its encoding mRNA, (b) proving the synthetic capacity of a cell in which a protein has been localized, (c) determining immunological or nucleic acid probe specificity, or (d) as an alternative to double-labeling immunocytochemical methods.

3,3'-Diaminobenzidine↗
PubMed1986

In situ hybridization technique to localize rRNA and mRNA in mammalian neurons.

In situ hybridization provides a method for identifying cells that contain specific nucleic acid sequences. This report outlines an in situ hybridization procedure for mammalian neural tissue. The method maintains morphological quality and produces excellent specificity. Seven tritiated nucleic acid probes were examined: two ribosomal RNA probes, a control pBR322 plasmid probe, two probes encoding portions of the gene for oxytocin, one probe each encoding a portion of vasopressin glycoprotein, and neurophysin. Using cryostat-cut rat brain sections, rRNA probes labeled the cytoplasm of all cells and the nucleoli of larger neurons. The plasmid probe failed to produce a strong signal. Oxytocin and vasopressin probes appropriately labeled the cytoplasm of hypothalamic magnocellular neurons. Vasopressin parvocellular neurons were not identified by the current method, and the shorter length neurophysin probe failed to produce a signal. Methodological variables were examined by counting autoradiographic grains in cells. The longer oxytocin probe produced a stronger signal than the shorter oxytocin and vasopressin probes, and higher probe concentrations resulted in stronger signal. Hybridization could be abolished by tissue pretreatment with RNAse A, and longer exposure time increased signal strength. The outlined fixation steps with fresh-frozen tissue produced a superior signal compared to paraformaldehyde-perfused tissue.

Animals↗
PubMed1985

An autoradiographic study of projections ascending from the midbrain central gray, and from the region lateral to it, in the rat.

Ascending projections from the midbrain central gray (CG) and from the region lateral to it were traced in the rat using tritiated amino acid autoradiography. Leucine or a cocktail of amino acids (leucine, proline, lysine, histidine, and tyrosine) were used as tracers. In addition to projections within the midbrain, ascending fibers follow three trajectories. The ventral projection passes through the ventral tegmental region of Tsai and the medial forebrain bundle to reach the hypothalamus, preoptic area, caudoputamen, substantia innominata, stria terminalis, and amygdala. There are labeled fibers in the diagonal bands of Broca and medial septum, and terminal labeling in the lateral septum, nucleus accumbens, olfactory tubercle, and frontal cortex. The dorsal periventricular projection terminates in the midline and intralaminar thalamic nuclei. The ventral periventricular projection follows the ventral component of the third ventricle into the hypothalamus, passing primarily through the dorsal hypothalamic area and labeling the rostral hypothalamus and preoptic area. Projections from the region lateral to the CG are similar, but exhibit stronger proximal, and weaker distal, projections. Rostral levels of the CG send heavier projections to the fields of Forel and the zona incerta, but fewer fibers through the supraoptic decussation, than do caudal levels. Ascending projections from the CG are both strong and widespread. Strong projections to the limbic system and the intralaminar thalamic nuclei provide an anatomical substrate for CG involvement in nociception and affective responses.

Amino Acids↗
PubMed1985

Estrogen effects on neuronal responsiveness to electrical and neurotransmitter stimulation: an in vitro study on the ventromedial nucleus of the hypothalamus.

To investigate the effects of estrogen on the activity of the neurons in the hypothalamic ventromedial nucleus (VMN), the single-unit activity was recorded from in vitro brain tissue slices prepared from ovariectomized rats that were either treated or untreated with estrogen. In tissue slices continuously perifused with estrogen-free solution, half of the 348 VMN neurons studied were silent or nearly silent, and the rest fired spontaneously at low rates. Electrical stimulation of the periventricular region medial to the VMN evoked not only 3 types of orthodromic responses, but also antidromic responses from many VMN neurons, indicating that the VMN and the periventricular region are intimately and reciprocally connected. Neuronal responses to a battery of 6 neurotransmitters were studied by microinjecting 50 microliters of a transmitter solution directly into the chamber where the tissue slice was being continuously perifused. When effective, glutamate (Glut) was almost exclusively excitatory; acetylcholine (ACh) and norepinephrine (NE) could be excitatory and/or inhibitory; 5-hydroxytryptamine (5-HT) and dopamine (DA) were predominantly inhibitory; and gamma-amino-butyric acid (GABA) was exclusively inhibitory on neuronal activity. The overall resting neuronal activity and responses to the transmitter were very similar to those observed in in vivo studies, indicating that, for VMN at least, in vitro tissue slices can serve as a simplified model for studying neuronal activity and responses. Comparisons of results between estrogen-treated and untreated preparations showed that chronic, in vivo estrogen treatments, while having no effect on resting firing rate, selectively facilitated the responsiveness of silent (but not spontaneously active) neurons to excitatory orthodromic inputs, and the responsiveness to the excitatory action of ACh, the overall action of NE and the inhibitory action of 5-HT. These selective estrogen effects would no doubt alter the input-output relationships of VMN neurons evoked by the orthodromic inputs and ACh, NE and 5-HT. Since the VMN is crucial to many biological functions, and the cholinergic, adrenergic and serotonergic inputs to the ventromedial hypothalamus are involved in the estrogenic regulation of sexual behavior, reproductive neuroendocrine, and other biological functions, the alteration of the input-output relationships evoked by the orthodromic and transmitter inputs would affect the regulation of many functions.

Acetylcholine↗
PubMed1985

Early estrogen-induced nuclear changes in rat hypothalamic ventromedial neurons: an ultrastructural and morphometric analysis.

Alterations in the fine structure of nuclei of ventromedial hypothalamic neurons in ovariectomized (OVX) rats after either a 2-hour exposure to estradiol (E2) or a discontinuous exposure (2 hours E2/7 hours off/2 hours E2), previously shown sufficient for female rat sexual behavior (Parsons et al., '82a), were examined with the electron microscope. Morphometric measurements of nucleolar, nuclear, and somal areas, and nuclear shape and perimeter were accomplished at the light microscope level. After 2 hours of E2, the appearance of the nucleoplasm was altered, with a decrease in the small, scattered clumps of heterochromatin. Nuclear shape appeared dramatically altered from nonspherical, invaginated profiles toward spherical, smooth profiles. Nucleolar, nuclear, and somal hypertrophy were evident. In addition, stacked rough endoplasmic reticulum was present more frequently in E2-treated than control OVX neurons. After the discontinuous (2 hours/7 hours/2 hours) E2 treatment, progressive loss of small clumps of heterochromatin along the nuclear envelope as well as in the nucleoplasm had occurred. Decrease in a large heterochromatin clump along the nuclear envelope was correlated with an increase in nucleolus-associated chromatin. As determined by a distribution analysis, these estrogen-induced nuclear changes co-occurred more frequently than predicted from mutual independence. These findings, the marked co-occurrence of E2-induced changes in 30% or more of the cells, and the differences between the 2-hour E2 and the 2-hour/7-hour/2-hour group are consistent with a cascade of cell nuclear changes in the first few hours after estrogen onset.

Animals↗
PubMed1985

Inhibition of the lordosis reflex in rats by intrahypothalamic infusion of neural excitatory agents: evidence that the hypothalamus contains separate inhibitory and facilitatory elements.

In attempts to activate lordosis-facilitating neural mechanisms in the ventromedial hypothalamus (VMH), neural excitatory agents were infused into the medial hypothalamus, and the effects of the infusions on the lordosis reflex and on the electrical activity of VMH neurons were studied. Surprisingly, bilateral intrahypothalamic infusion of glutamate (10 mM, 1.0 microliter/side) into behaving, ovariectomized, estrogen-treated rats displaying moderate lordotic responsiveness did not facilitate lordosis, but instead, resulted in a rapid (within a few minutes) and transient (recovery in about 20 min) inhibition of lordosis. Further experiments showed that this lordosis-inhibiting effect of glutamate was dose-related, and was completely blocked by prior infusion of a local anesthetic, procaine. Infusion of KC1 (1.5 or 15%, 1.0 microliter/side) also induced a dose-related, rapid and transient inhibition of lordosis, that was essentially identical to that induced by glutamate. Kainic acid (0.25 micrograms/0.5 microliter/side) also caused a rapid inhibition of lordosis, but the effect was long-lasting (days). The inhibition of lordosis by these agents was dissociated in time course, presence, and/or severity from effects on non-lordotic behaviors. Electrophysiological studies showed that all three agents tested could excite multiunit activity of the VMH, and that the time courses of these excitations were closely comparable to those of the inhibition of lordosis induced by the respective agents. Altogether, these studies indicated that the excitation of certain medial hypothalamic neurons can inhibit the lordosis reflex. The implied lordosis-inhibiting neural mechanisms are separate from facilitatory mechanism(s), according to differences in latency, duration, and procaine-sensitivity of response.

Animals↗
PubMed1985

Ontogenesis of neurons producing luteinizing hormone-releasing hormone (LHRH) in the nervus terminalis of the rat.

Immunoreactive luteinizing hormone-releasing hormone (LHRH) was first detected at 15 days of gestation in ganglion cells associated with the peripheral, intracranial, and central parts of the nervus terminalis of the rat. LHRH was not detected in any other structure of the central nervous system at this age. In the 17-day-old fetal rat, 62% of the total LHRH-reactive neuronal population was found in ganglion cells of the nervus terminalis. At this same age, immunoreactive beta-luteinizing hormone (beta-LH) was first seen in gonadotropes of the anterior pituitary gland. At 19 days of gestation, 31% of the total number of LHRH-reactive neurons observed in the rat brain was found in the nervus terminalis, and immunoreactive processes were first seen in the organum vasculosum of the lamina terminalis and in the median eminence. Our data indicate that from 15 to 19 days of gestation the nervus terminalis is a principal source of LHRH in the fetal rat. Presence of the decapeptide in the nervus terminalis prior to appearance of beta-LH in the anterior pituitary suggests a possible role for LHRH in this system on maturation of the gonadotropes and differentiation of the brain-pituitary-gonadal axis.

Animals↗
PubMed1985

Anatomical identification of neurons in selected brain regions associated with maternal behavior deficits induced by knife cuts of the lateral hypothalamus in rats.

The present experiment identified neurons associated with maternal behavior deficits induced by damage to the lateral hypothalamus (LH) in rats. Fully maternal lactating rats received bilateral coronal knife cuts through either the dorsal or ventral LH at the level of the ventromedial nucleus. The blade of the wire knife used to make the cuts was coated with horseradish peroxidase (HRP). The maternal behavior of all females was studied for 4 days postoperatively and then the brains were processed for the localization of neurons retrogradely filled with HRP. The analysis focused on those brain regions thought to be important for maternal behavior. The dorsal LH cuts severely disrupted maternal behavior while the ventral LH cuts did not. The ventral cuts labeled more medial preoptic area (MPOA) and septal-diagonal band neurons with HRP than did the dorsal cuts. The dorsal LH cuts labeled more neurons with HRP in the lateral preoptic area (LPOA), magnocellular preoptic area, bed nucleus of the stria terminalis, ventral tegmental area (VTA), substantia nigra, and central gray than did the ventral cuts. Previous research has suggested that the lateral efferents of the MPOA interact with the VTA in the control of maternal behavior. The results of the present experiment suggest that MPOA axons which descend directly to the brainstem via the ventral LH are not essential for maternal behavior. Our results are consistent with the view that an MPOA-to-LPOA-to-VTA circuit may be important for maternal behavior.

Animals↗
PubMed1985

The pudendal nerve-evoked response in axial muscle.

In 39 Urethane-anesthetized rats we have recorded the afferent volley in the dorsal roots and the electrical activity of the lateral longissimus muscle and its motor nerves during electrical stimulation of a cutaneous branch of the pudendal nerve. Male and female rats were used; the females were ovariectomized and either pretreated with estradiol or left without hormonal treatment. Conduction velocities in the pudendal nerve were 54 m/s for the largest A-beta fibers and averaged 10 m/s for A-delta fibers. Excitation of pudendal nerve afferents strongly potentiated the firing of axial motoneurons, at stimulus currents below threshold for A-delta fibers. Trains of three shocks to the pudendal nerve were considerably more effective than double or single shock trains. Repetition rates as low as 1/s had a long lasting excitatory effect on the lateral longissimus muscle and the magnitude of the responses increased gradually for several seconds with continued stimulation. Recordings from the axons of the epaxial motoneurons of female rats showed a strong activation of neuronal firing with an onset latency of 5.8 ms from the last shock of a three ms, three shock train; the onset in male rats, 8.4 ms, differed significantly. Peak spike activity occurred at mean latencies of 11, 22 and 102 ms in both sexes. A period of depressed firing was usually present from 34 to 50 ms. Males differed in having a larger peak in activity at 102 ms, but the overall profile of the responses was similar in males and females. No differences were seen in the overall response patterns of the estrogen-treated and untreated females. Responses of comparable magnitude were seen with ipsilateral or contralateral pudendal nerve stimulation; these were facilitated by bilateral stimulation. In electromyographic recordings, the onset of unit firings was seen at 6.4 ms latency in response to pudendal nerve stimulation. This unit activity was consistent with the firing pattern seen in the muscle nerves.

Animals↗
PubMed1985

Immunocytochemical localization of actin in dendritic spines of the cerebral cortex using colloidal gold as a probe.

Immunocytochemical localization of actin in rat cerebral cortex embedded in the resin LR White was performed using 5 nm colloidal gold as a probe. Antigenicity is maintained throughout the embedding procedure and the low electron opacity of LR White permits fine filamentous structures to be visualized. Control experiments included incubating the sections with normal goat serum or mouse IgG instead of the primary antibody, preadsorbing the antibody with actin from bovine muscle or liver acetone powder, and heat treating the primary antibody. Immunoreactive actin was identified primarily in dendritic spines, particularly in the postsynaptic density (PSD), the subsynaptic web, and the spine apparatus and endothelial and smooth muscle cells of blood vessels. Within dendritic spines, actin which is labeled in the PSD is in continuity with the filaments of the subsynaptic web. These filaments, in turn, are in continuity with the spine apparatus and/or the spine membranes adjacent to the PSD. The PSD may therefore function like other submembranous filamentous arrays which communicate events occurring at the membrane, in this case, the postsynaptic membrane, to the underlying cytoskeletal network, i.e., the subsynaptic web of the spine. It is also suggested that the actin present in the spine may play a role in changes in spine shape and synaptic curvature. Some actin was also seen in the presynaptic process in association with synaptic vesicles, the filamentous network that is contiguous with the synaptic vesicle membrane, and the presynaptic dense projections. Actin may be involved in dynamic processes in the presynaptic ending which include vesicle translocation.

Actins↗
PubMed1985

Specificity and neural sites of action of anisomycin in the reduction or facilitation of female sexual behavior in rats.

These experiments were designed to investigate the role of neuronal protein synthesis in the hormonal activation of female sexual behavior using intracranial implants of the protein synthesis inhibitor, anisomycin. In the first experiment, female rats receiving bilateral cannulae implants in the medial preoptic area (POA), septal region (SEPT), ventromedial hypothalamus (VMH), or midbrain central gray (CG) were injected with 2.5 micrograms estradiol benzoate (EB), followed 48 hr later by 500 micrograms progesterone (P). Females receiving anisomycin in the VMH at the time of EB injection had lower levels of lordosis and darting compared to tests without anisomycin. Sexual behavior was unaffected in females receiving anisomycin implants in the POA, SEPT, or CG. In a second experiment, we replicated the finding that anisomycin could attenuate lordotic responsivity when placed in the VMH of female rats injected with 2.5 micrograms EB and 500 micrograms P. In addition, we found that POA implants of anisomycin could facilitate lordosis in females given a low dose of EB (1.25 microgram) plus 500 micrograms P. In a third experiment, we assessed the effects of anisomycin application to the VMH or POA of female rats receiving estradiol (E; diluted 1:250 with cholesterol) implants in the VMH and systemic P. Treatment of the VMH with anisomycin prior to E in the VMH suppressed lordotic responding, whereas anisomycin application to the POA prior to E in the VMH had no effect on lordosis. The results of these experiments suggest that reducing protein synthesis in the region of the VMH disrupts the action of estrogen on the VMH, and that the facilitative action of anisomycin in the POA of female rats requires more estrogen treatment than threshold stimulation of the VMH alone.

Animals↗
PubMed1985

Preoptic implants of estradiol increase wheel running but not the open field activity of female rats.

In order to localize the neural substrate for estrogen induction of increases in activity, bilateral implants of dilute estradiol (in concentrations of 1:10, 1:100, and 1:250) were made into the medial preoptic area (n = 5), the anterior hypothalamus (n = 8), the ventromedial hypothalamus (n = 6), and the posterior hypothalamus (n = 4) of ovariectomized female rats housed in running wheels. The same animals were also tested for open field activity and lordosis. The preoptic area was the most effective site tested for estrogenic stimulation of running wheel activity, with 3/5 animals responding consistently to estradiol application. 3/8 animals with cannulae in the anterior hypothalamus (AHA) responded with increases in activity measured in the running wheel on at least two out of three estradiol treatments. No animals with cannula placements posterior to the AHA exhibited consistent running wheel responses to estradiol. No estrogen-induced increase in open field activity was found, regardless of cannula location or concentration of estradiol; however, there was a tendency for the number of squares entered during the later tests to decrease in the ventromedial hypothalamic nucleus group and in the subgroup of animals not showing estrogen-sensitive wheel running in the anterior hypothalamic area group. Only the ventromedial hypothalamic estradiol treatments stimulated lordosis behavior. These results support earlier reports suggesting that the medial preoptic area (and possibly the adjacent anterior hypothalamic area) is the critical brain site for estrogenic stimulation of running wheel activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Brain region specificity in estradiol effects on neuronal ultrastructure in rats.

The effects of long-term (15 days) and short-term (3 days) treatment with estradiol benzoate (EB) on stacking of rough endoplasmic reticulum in neurons of four brain regions (medial preoptic nucleus, ventromedial hypothalamus, arcuate nucleus and midbrain central gray) were investigated. These regions were chosen for their role in reproduction in female rats and/or the presence of estradiol-concentrating cells. Long-term EB treatment increased the proportion of neurons in the ventromedial hypothalamus (by 30%) and medial preoptic nucleus (by 17%) that contained stacks of rough endoplasmic reticulum. Short-term EB affected stacking only in ventromedial hypothalamic neurons. EB was without effect on neurons of the arcuate nucleus of the hypothalamus or midbrain central gray. The degree of stacking of rough endoplasmic reticulum in neurons of the ventromedial hypothalamus was positively related to the level of sexual behavior (lordosis) activated by these hormone treatments.

Animals↗