A hypothalamic-thalamostriatal circuit that controls approach-avoidance conflict in rats.

Survival depends on a balance between seeking rewards and avoiding potential threats, but the neural circuits that regulate this motivational conflict remain largely unknown. Using an approach-food vs. avoid-predator threat conflict test in rats, we identified a subpopulation of neurons in the anterior portion of the paraventricular thalamic nucleus (aPVT) which express corticotrophin-releasing factor (CRF) and are preferentially recruited during conflict. Inactivation of aPVTCRF neurons during conflict biases animal's response toward food, whereas activation of these cells recapitulates the food-seeking suppression observed during conflict. aPVTCRF neurons project densely to the nucleus accumbens (NAc), and activity in this pathway reduces food seeking and increases avoidance. In addition, we identified the ventromedial hypothalamus (VMH) as a critical input to aPVTCRF neurons, and demonstrated that VMH-aPVT neurons mediate defensive behaviors exclusively during conflict. Together, our findings describe a hypothalamic-thalamostriatal circuit that suppresses reward-seeking behavior under the competing demands of avoiding threats.

Disruption of reproductive aging in female and male rats by gestational exposure to estrogenic endocrine disruptors.

Polychlorinated biphenyls (PCBs) are industrial contaminants and known endocrine-disrupting chemicals. Previous work has shown that gestational exposure to PCBs cause changes in reproductive neuroendocrine processes. Here we extended work farther down the life spectrum and tested the hypothesis that early life exposure to Aroclor 1221 (A1221), a mixture of primarily estrogenic PCBs, results in sexually dimorphic aging-associated alterations to reproductive parameters in rats, and gene expression changes in hypothalamic nuclei that regulate reproductive function. Pregnant Sprague Dawley rats were injected on gestational days 16 and 18 with vehicle (dimethylsulfoxide), A1221 (1 mg/kg), or estradiol benzoate (50 µg/kg). Developmental parameters, estrous cyclicity (females), and timing of reproductive senescence were monitored in the offspring through 9 months of age. Expression of 48 genes was measured in 3 hypothalamic nuclei: the anteroventral periventricular nucleus (AVPV), arcuate nucleus (ARC), and median eminence (females only) by real-time RT-PCR. Serum LH, testosterone, and estradiol were assayed in the same animals. In males, A1221 had no effects; however, prenatal estradiol benzoate increased serum estradiol, gene expression in the AVPV (1 gene), and ARC (2 genes) compared with controls. In females, estrous cycles were longer in the A1221-exposed females throughout the life cycle. Gene expression was not affected in the AVPV, but significant changes were caused by A1221 in the ARC and median eminence as a function of cycling status. Bionetwork analysis demonstrated fundamental differences in physiology and gene expression between cycling and acyclic females independent of treatment. Thus, gestational exposure to biologically relevant levels of estrogenic endocrine-disrupting chemicals has sexually dimorphic effects, with an altered transition to reproductive aging in female rats but relatively little effect in males.

Neonatal exposure to genistein adversely impacts the ontogeny of hypothalamic kisspeptin signaling pathways and ovarian development in the peripubertal female rat.

Neonatal exposure to estrogenic endocrine disrupting compounds (EDCs) can advance pubertal onset and induce premature anestrous in female rats. It was recently discovered that hypothalamic kisspeptin (KISS) signaling pathways are sexually dimorphic and regulate both the timing of pubertal onset and estrous cyclicity. Thus we hypothesized that disrupted sex specific ontogeny of KISS signaling pathways might be a mechanism underlying these EDC effects. We first established the sex specific development of KISS gene expression, cell number and neural fiber density across peripuberty in the anteroventral periventricular nucleus (AVPV) and arcuate nucleus (ARC), hypothesizing that the sexually dimorphic aspects of KISS signaling would be most vulnerable to EDCs. We next exposed female rats to the phytoestrogen genistein (GEN, 1 or 10 mg/kg bw), estradiol benzoate (EB, 10 µg), or vehicle from post natal day (P) 0-3 via subcutaneous (sc) injection. Animals were sacrificed on either P21, 24, 28, or 33 (n=5-14 per group at each age). Vaginal opening was significantly advanced by EB and the higher dose of GEN compared to control animals and was accompanied by lower numbers of KISS immunoreactive fibers in the AVPV and ARC. Ovarian morphology was also assessed in all age groups for the presence of multiple oocyte follicles (MOFs). The number of MOFs decreased over time in each group, and none were observed in control animals by P24. MOFs were still present, however, in the EB and 10 mg/kg GEN groups beyond P24 indicating a disruption in the timing of ovarian development.

The targets of acetone cyanohydrin neurotoxicity in the rat are not the ones expected in an animal model of konzo.

Konzo is a neurotoxic motor disease caused by excess consumption of insufficiently processed cassava. Cassava contains the cyanogenic glucoside linamarin, but konzo does not present the known pathological effects of cyanide. We hypothesized that the aglycone of linamarin, acetone cyanohydrin, may be the cause of konzo. This nitrile rapidly decomposes into cyanide and acetone, but the particular exposure and nutrition conditions involved in the emergence of konzo may favor its stabilization and subsequent acute neurotoxicity. A number of preliminary observations were used to design an experiment to test this hypothesis. In the experiment, young female Long-Evans rats were given 10mM acetone cyanohydrin in drinking water for 2 weeks, and then 20mM for 6 weeks. Nutrition deficits associated with konzo were modeled by providing tapioca (cassava starch) as food for the last 3 of these weeks. After this period, rats were fasted for 24h in order to increase endogenous acetone synthesis, and then exposed to 0 (control group) or 50 micromol/kg-h of acetone cyanohydrin for 24h (treated group) through subcutaneous osmotic minipump infusion (n=6/group). Motor activity and gait were evaluated before exposure (pre-test), and 1 and 6 days after exposure. Brains (n=4) were stained for neuronal degeneration by fluoro-jade B. Rats exposed to 50 micromol/kg-h of acetone cyanohydrin showed acute signs of toxicity, but no persistent motor deficits. Two animals showed fluoro-jade staining in discrete thalamic nuclei, including the paraventricular and the ventral reuniens nuclei; one also exhibited labeling of the dorsal endopiriform nucleus. Similar effects were not elicited by equimolar KCN exposure. Therefore, acetone cyanohydrin may cause selective neuronal degeneration in the rat, but the affected areas are not those expected in an animal model of konzo.

Effect of reversible inactivation of the reuniens nucleus on spatial learning and memory in rats using Morris water maze task.

The involvement of thalamic midline nuclei (MLN) in early stage of Alzheimer's disease and in diencephalic amnesia has drawn attention to the connectivity between the nucleus reuniens (RE) and structures of medial temporal lobe. RE is major source of thalamic afferents to the hippocampus and has been shown to exert powerful excitatory action on CA1 of hippocampus, which is supposed to be involved in learning and memory processes. However, the role of the RE on spatial memory is a controversial issue. The present study was designed to evaluate the role of the RE in acquisition, consolidation and retrieval of spatial reference memory (RM) and working memory (WM). We assessed the effect of reversible inactivation of RE with tetracaine (0.5 microl, 2%) on different stages of memory. Rats were trained on RM and WM versions of the Morris water maze (MWM) task. RE was inactivated before or immediately after training or before the probe trial of retrieval tests. The data showed that reversible inactivation of the RE significantly impaired both RM and WM versions of MWM. Therefore, it seems that nucleus reuniens of thalamus plays a role in spatial RM and WM version of the MWM task in rats.

Activation of orexin/hypocretin projections to basal forebrain and paraventricular thalamus by acute nicotine.

Orexin/hypocretin neurons of the lateral hypothalamus/perifornical area project to a diverse array of brain regions and are responsive to a variety of psychostimulant drugs. It has been shown that orexin neurons are activated by systemic nicotine administration suggesting a possible orexinergic contribution to the effects of this drug on arousal and cognitive function. The basal forebrain and paraventricular nucleus of the dorsal thalamus (PVT) both receive orexin inputs and have been implicated in arousal, attention and psychostimulant drug responses. However, it is unknown whether orexin inputs to these areas are activated by psychostimulant drugs such as nicotine. Here, we infused the retrograde tract tracer cholera toxin B subunit (CTb) into either the basal forebrain or PVT of adult male rats. Seven to 10 days later, animals received an acute systemic administration of (-) nicotine hydrogen tartrate or vehicle and were euthanized 2h later. Triple-label immunohistochemistry/immunofluorescence was used to detect Fos expression in retrogradely-labeled orexin neurons. Nicotine increased Fos expression in orexin neurons projecting to both basal forebrain and PVT. The relative activation in lateral and medial banks of retrogradely-labeled orexin neurons was similar following basal forebrain CTb deposits, but was more pronounced in the medial bank following PVT deposits of CTb. Our findings suggest that orexin inputs to the basal forebrain and PVT may contribute to nicotine effects on arousal and cognition and provide further support for the existence of functional heterogeneity across the medial-lateral distribution of orexin neurons.

Medial auditory thalamus inactivation prevents acquisition and retention of eyeblink conditioning.

The auditory conditioned stimulus (CS) pathway that is necessary for delay eyeblink conditioning was investigated using reversible inactivation of the medial auditory thalamic nuclei (MATN) consisting of the medial division of the medial geniculate (MGm), suprageniculate (SG), and posterior intralaminar nucleus (PIN). Rats were given saline or muscimol infusions into the MATN contralateral to the trained eye before each of four conditioning sessions with an auditory CS. Rats were then given four additional sessions without infusions to assess savings from the initial training. All rats were then given a retention test with a muscimol infusion followed by a recovery session. Muscimol infusions through cannula placements within 0.5 mm of the MGm prevented acquisition of eyeblink conditioned responses (CRs) and also blocked CR retention. Cannula placements more than 0.5 mm from the MATN did not completely block CR acquisition and had a partial effect on CR retention. The primary and secondary effects of MATN inactivation were examined with 2-deoxy-glucose (2-DG) autoradiography. Differences in 2-DG uptake in the auditory thalamus were consistent with the cannula placements and behavioral results. Differences in 2-DG uptake were found between groups in the ipsilateral auditory cortex, basilar pontine nuclei, and inferior colliculus. Results from this experiment indicate that the MATN contralateral to the trained eye and its projection to the pontine nuclei are necessary for acquisition and retention of eyeblink CRs to an auditory CS.

Severe reversible toxic encephalopathy induced by cisplatin in a patient with cervical carcinoma receiving combined radiochemotherapy.

CASE REPORT: A 45-year-old patient with cervix carcinoma received combined radiochemotherapy including cisplatin. After a cumulative dose of 240 mg/m(2) the patient suddenly became somnolent and developed a severe tetraparesis and generalized seizures. After ruling out intracranial bleeding, cerebral metastases as well as infectious and metabolic causes of this condition, a severe toxic encephalopathy was diagnosed based on the clinical findings and MRI scans. After symptomatic treatment on the intensive care unit all symptoms were completely reversible. CONCLUSION: Toxic encephalopathy is a rare but dramatic complication of various cytostatic drugs. With the widespread use of cisplatin this rare disorder should be kept in mind.

AVPV neurons containing estrogen receptor-beta in adult male rats are influenced by soy isoflavones.

BACKGROUND: Isoflavones, the most abundant phytoestrogens in soy foods, are structurally similar to 17beta-estradiol. It is known that 17beta-estradiol induces apoptosis in anteroventral periventricular nucleus (AVPV) in rat brain. Also, there is evidence that consumption of soy isoflavones reduces the volume of AVPV in male rats. Therefore, in this study, we examined the influence of dietary soy isoflavones on apoptosis in AVPV of 150 day-old male rats fed either a soy isoflavone-free diet (Phyto-free) or a soy isoflavone-rich diet (Phyto-600). RESULTS: The occurrence of apoptosis in AVPV was examined by TUNEL staining. The incidence of apoptosis was about 10 times higher in the Phyto-600 group (33.1 +/- 1.7%) than in the Phyto-free group (3.6 +/- 1.0%). Furthermore, these apoptotic cells were identified as neurons by dual immunofluorescent staining of GFAP and NeuN as markers of astrocytes and neurons, respectively. Then the dopaminergic neurons in AVPV were detected by immunohistochemistry staining of tyrosine hydroxylase (TH). No significant difference in the number of TH neurons was observed between the diet treatment groups. When estrogen receptor (ER) alpha and beta were examined by immunohistochemistry, we observed a 22% reduction of ERbeta-positive cell numbers in AVPV with consumption of soy isoflavones, whereas no significant change in ERalpha-positive cell numbers was detected. Furthermore, almost all the apoptotic cells were ERbeta-immunoreactive (ir), but not ERalpha-ir. Last, subcutaneous injections of equol (a major isoflavone metabolite) that accounts for approximately 70-90% of the total circulating plasma isoflavone levels did not alter the volume of AVPV in adult male rats. CONCLUSION: In summary, these findings provide direct evidence that consumption of soy isoflavones, but not the exposure to equol, influences the loss of ERbeta-containing neurons in male AVPV.

Innervation of the paraventricular nucleus of the thalamus from cocaine- and amphetamine-regulated transcript (CART) containing neurons of the hypothalamus.

The paraventricular nucleus of the thalamus (PVT) is a midline thalamic nucleus with heavy projections to the nucleus accumbens and other limbic regions. Previous studies have shown that the PVT contains fibers immunoreactive for cocaine- and amphetamine-related transcript (CART). The purpose of the present study was to determine the location of CART neurons innervating the PVT of the rat by using retrograde tracing with cholera toxin B (CTb) combined with immunofluorescence for CTb and CART (amino acid sequence 55-102). Immunohistochemical analysis of CART in the dorsal thalamus showed that the PVT is densely innervated by CART fibers whereas adjacent midline and intralaminar thalamic nuclei are unlabeled. Injections of CTb in the dorsal midline thalamus retrogradely labeled neurons in several areas of the hypothalamus and brainstem which also contained CART neurons. The largest number of double-labeled neurons (CTb/CART) was found in the arcuate nucleus of the hypothalamus. CTb/CART neurons were also found in the lateral hypothalamus, zona incerta, and periventricular hypothalamus. These results indicate that the arcuate nucleus is a major source of CART fibers in the PVT. CART neurons in the arcuate nucleus monitor circulating hormonal signals and may regulate food intake and hypothalamic-pituitary-adrenal (HPA) activity. Consequently, CART neurons in the arcuate nucleus may transmit signals to the PVT which in turn may influence limbic regions involved in regulating food intake and the HPA.

Central relaxin-3 administration causes hyperphagia in male Wistar rats.

Relaxin-3 (INSL-7) is a recently discovered member of the insulin superfamily. Relaxin-3 mRNA is expressed in the nucleus incertus of the brainstem, which has projections to the hypothalamus. Relaxin-3 binds with high affinity to the LGR7 receptor and to the previously orphan G protein-coupled receptor GPCR135. GPCR135 mRNA is expressed predominantly in the central nervous system, particularly in the paraventricular nucleus (PVN). The presence of relaxin-3 and these receptors in the PVN led us to investigate the effect of central administration of relaxin-3 on food intake in male Wistar rats. The receptor involved in mediating these effects was also investigated. Intracerebroventricular injections of human relaxin-3 (H3) to satiated rats significantly increased food intake 1 h post administration in the early light phase [0.96 +/- 0.16 g (vehicle) vs. 1.81 +/- 0.21 g (180 pmol H3), P < 0.05] and the early dark phase [2.95 +/- 0.45 g (vehicle) vs. 4.39 +/- 0.39 g (180 pmol H3), P < 0.05]. Intra-PVN H3 administration significantly increased 1-h food intake in satiated rats in the early light phase [0.34 +/- 0.16 g (vehicle) vs. 1.23 +/- 0.30 g (18 pmol H3), P < 0.05] and the early dark phase [4.43 +/- 0.32 g (vehicle) vs. 6.57 +/- 0.42 g (18 pmol H3), P < 0.05]. Feeding behavior increased after intra-PVN H3. Equimolar doses of human relaxin-2, which binds the LGR7 receptor but not GPCR135, did not increase feeding. Hypothalamic neuropeptide Y, proopiomelanocortin, or agouti-related peptide mRNA expression did not change after acute intracerebroventricular H3. These results suggest a novel role for relaxin-3 in appetite regulation.

Estrogens and phytoestrogens: brain plasticity of sexually dimorphic brain volumes.

Sexually dimorphic brain volumes (sexually dimorphic nucleus of the preoptic area (SDN-POA) and anteroventral periventricular (AVPV) nucleus) are influenced by estrogens. Phytoestrogens, derived from plants (especially soy products), are molecules structurally and functionally similar to estradiol. The purpose of this study was to examine: the consumption of phytoestrogen (using a phytoestrogen-rich (Phyto-600) versus a phytoestrogen-free (Phyto-free)) diets from conception to adulthood (or changing the diets during adulthood) and characterizing (a) circulating plasma phytoestrogen levels, (b) testosterone levels in males, (c) sexually dimorphic brain volumes (i.e. the SDN-POA and AVPV) and (d) the presence of apoptotic cells in these brain structures in Long-Evans rats. Phyto-600 fed animals displayed total serum phytoestrogens levels 37-fold higher compared to Phyto-free values. Circulating testosterone levels were not significantly altered by the diets. Female SDN-POA volumes were not altered by the diets. Whereas, males fed a Phyto-free diet displayed decreased SDN-POA volumes compared to male Phyto-600 values. Females fed the Phyto-600 diet displayed larger AVPV volumes compared to males on the same diet or females on the Phyto-free diet. Males fed the Phyto-free diet had the largest AVPV values compared to Phyto-600 fed males. When the SDN-POA region was examined in lifelong Phyto-free fed males, apoptotic cells were present versus males fed the Phyto-600 diet and in the AVPV region the opposite results were obtained. In summary, consumption of dietary phytoestrogens (estrogen mimics) can alter hormone-sensitive hypothalamic brain volumes in rodents during adulthood.

Magnetic resonance imaging of changes elicited by status epilepticus in the rat brain: diffusion-weighted and T2-weighted images, regional blood volume maps, and direct correlation with tissue and cell damage.

The rat brain was investigated with structural and functional magnetic resonance imaging (MRI) 12 h after the arrest of pilocarpine-induced status epilepticus lasting 4 h. Histopathological data, obtained immediately after MRI analysis, were correlated with the images through careful evaluation of tissue shrinkage. Diffusion-weighted and T2-weighted imaging showed changes throughout the cerebral cortex, hippocampus, amygdala, and medial thalamus. However, only T2-weighted imaging, based on rapid acquisition relaxation-enhanced sequences, revealed in the cortex inhomogeneous hyperintensity that was highest in a band corresponding to layer V. Regional cerebral blood volume (rCBV) maps were generated using T2*-weighted gradient-echo images and an ultrasmall superparamagnetic iron oxide contrast agent. In the cortex, rCBV peaked in superficial and deep bands exhibiting a distribution complementary to the highest T2-weighted intensity. Selective rCBV increase was also documented in the hippocampus and subcortical structures. In tissue sections, alterations indicative of marked edema were found with Nissl staining in areas corresponding to the highest T2-weighted intensity. Degenerating neurons, revealed by FluoroJadeB histochemistry, were instead concentrated in tissue exhibiting hyperperfusion in rCBV maps, such as hippocampal subfields and dentate gyrus, cortical layers II/III and VI, and medial thalamus. The data indicate that:(i) T2-weighted imaging provides a sensitive tool to investigate edematous brain alterations that follow sustained seizures; (ii) rCBV maps reveal regional hyperperfusion; (iii) rCBV peaks in tissue exhibiting marked neurodegeneration, which may not be selectively revealed by structural MRI. The findings provide an interpretation of the brain response to sustained seizures revealed in vivo by different strategies of MRI analysis.

Hypothalamic imprinting by gonadal steroid hormones.

The results of more than four decades of research on different mammalian species have established that the brain, like the rest of the reproductive system, is esentially basically female. For the male to develop structural and functional characteristics typical of his species, his brain must be exposed to testicular hormones during a critical period, or critical periods, of development. As mammals, human beings are most likely subject to this process of the hormone-dependent sexual differentiation of the brain, but proving it will be difficult. Common sense ethics preclude experimental procedures such as castration of neonatal infants or exposing the female fetus to testosterone perinatally. Thus, scientists are restricted to the retrospective study of "Experiments of Nature." The results of such studies support to a degree a meaningful role of hormones in the development of the human brain. The concept of the sexual differentiation of brain structure and function has a potentially profound influence on clinical decisions with respect to sex assignment and clinical management of infants with ambiguous or poorly developed external genitalia. Because of the importance of a baby's sex in our culture, parents of such infants must be given consideration, but so should the infant whose hormonal environment prenatally may well have produced permanent changes in the structure and functional potential of his/her brain.