Excitatory M5 muscarinic acetylcholine receptors are expressed at the highest level in the brain, in the ventromedial hypothalamus (VMH). M5 knockout (KO) mice emit fewer ultrasonic vocalizations (USVs) during mating, and show 40-50% lesser morphine-induced locomotion as compared to wild-type (WT) mice. Following viral transfection of M5 muscarinic receptors in the ventral tegmentum (VTA), KO mice depict restored mating-induced USVs and enhanced morphine-induced locomotion. The VMH is important for motivational processes, such as, feeding and producing USVs in rats. With a Herpes simplex virus (HSV), the M5 receptor gene was transfected into the VMH of WT mice. M5 transfection into neurons of the VMH increased locomotion in mice injected with 10mg/kg morphine. When a D2-selective dopamine blocker was injected into the same mice, locomotion was drastically reduced. There were no significant differences in amount of USVs produced. VMH may exert its effects on morphine through a DA dependent VTA pathway.

Excitatory M5 muscarinic acetylcholine receptors are expressed at the highest level in the brain, in the ventromedial hypothalamus (VMH). M5 knockout (KO) mice emit fewer ultrasonic vocalizations (USVs) during mating, and show 40-50% lesser morphine-induced locomotion as compared to wild-type (WT) mice. Following viral transfection of M5 muscarinic receptors in the ventral tegmentum (VTA), KO mice depict restored mating-induced USVs and enhanced morphine-induced locomotion. The VMH is important for motivational processes, such as, feeding and producing USVs in rats. With a Herpes simplex virus (HSV), the M5 receptor gene was transfected into the VMH of WT mice. M5 transfection into neurons of the VMH increased locomotion in mice injected with 10mg/kg morphine. When a D2-selective dopamine blocker was injected into the same mice, locomotion was drastically reduced. There were no significant differences in amount of USVs produced. VMH may exert its effects on morphine through a DA dependent VTA pathway.

Lesions of the ventromedial hypothalamus (VMH) result in an obesity syndrome characterized by metabolic and behavioral disturbances. It has recently been indicated that damage to the paraventricular nucleus of the hypothalamus (PVH) also leads to changes characteristic of obesity. Although deficits following VMH lesions have been characterized extensively, less is known about the consequences of PVH damage. This thesis presents a series of experiments providing a detailed comparison of the two hypothalamic lesion syndromes. Initially, to assess the basic features of the syndromes, rats underwent VMH, PVH, or sham lesions and were maintained ad libitum for 15 weeks on a series of test diets. Overall, lesion groups gained similar amounts of weight (significantly more than controls) and were equally hyperphagic. However, carcass analyses revealed that although both lesion groups had larger body fat compartments than controls, VMH rats were fatter than PVH animals. Similarly, although insulin levels in both lesion groups were elevated, only VMH rats had a significant hyperinsulinemia. A defining feature of the VMH obesity is the development of certain disturbances of visceral secretion and excessive adipose stores even in the absence of hyperphagia. To assess whether the PVH obesity shares these characteristics, food intake of PVH and VMH rats were restricted postlesion to control body weights. First, gastric acid secretion was measured to index lesion-induced changes in visceral secretion. VMH rats developed a persisting hypersecretion immediately postlesion; acid secretion levels of PVH rats were normal. In a second experiment, PVH and VMH rats were maintained at control weights for 28 days postlesion by restricted feeding of either a standard pellet or high fat diet. In both diet conditions, VMH rats became obese but PVH rats did not. Finally, the effects of PVH and VMH lesions on behavioral reactivity to orosensory properties of food were assessed by comparing the sham feeding responses of PVH, VMH and control rats to liquid diets varying in palatability. Control animals increased sham feeding with ascending sucrose concentrations. VMH animals showed disproportionately large increases in consumption with increased sucrose. PVH animals showed sham feeding changes similar to VMH rats. These data indicate similar effects of PVH and VMH lesions on behavioral measures; specifically, in normal, and sham, feeding. However, these two lesions produce different effects on metabolic and secretion measures. It is concluded that the etiologies of the two obesity are fundamentally different. / Thesis / Doctor of Philosophy (PhD)

Electrolytic lesions of the ventromedial hypothalamus produce an obesity syndrome in experimental animals characterized by behavioural and metabolic disturbances. Historically, theories of VMH obesity have considered a single disturbance, either behavioural or metabolic, to be the primary effect of the lesion, which in turn causes other components of the syndrome. An alternative view suggests that VMH lesions simultaneously disturb both behavioural and metabolic mechanisms due to the anatomical proximity of these mechanisms in the hypothalamus. Therefore, more discrete lesions in the VMH may produce some syndrome components but not others. This thesis presents a series of experiments that test this "dissociative" perspective of the VMH obesity syndrome. First, rats having different hypothalamic ablations were compared on: caloric intakes on a series of test diets, body weight changes, and body fat. Bilateral parafornical hypothalamic knife cuts (PFKC) that spared the ventromedial hypothalamic nucleus (VMN), produced overeating and weight gain characteristic of VMH lesions. However, measurement of percentage body fat (i.e. level of obesity) indicated that PFKC rats were less obese than VMH rats, even though PFKC lesions produced a greater hyperphagia and weight gain than VMH lesions. In contrast, lesions restricted to VMN produced obesity, but did not produce hyperphagia or weight gain. Since parafornical knife cuts produced a greater hyperphagia than VMH lesions, it is possible that VMN damage actually reduces caloric intake in VMH rats. To test this hypothesis, the effects of VMH, PFKC, and combined PFKC/VMN lesions on caloric intake and body weight were compared. PFKC and VMH lesions produced hyperphagia and weight gain. However, knife cuts were not significantly more effective than VMH lesions for producing these disturbances in this experiment. Therefore, PFKC lesions do not invariably produce a greater hyperphagia than VMH lesions. Furthermore, VMN lesions had no effect on the level of overeating or weight gain in rats bearing PFKC lesions. Therefore, damage to VMN does not reduce the hyperphagia produced by PFKC lesions. Finally, the effects of these different hypothalamic manipulations on metabolic measures were determined. To eliminate the confound of hyperphagia on metabolic variables, all lesion rats were fed a daily food ration sufficient to maintain their body weight at the level of controls. VMH and PFKC lesions resulted in elevated parasympathetic tone, indicated by elevated basal gastric acid secretion. VMN lesions did not affect gastric acid secretion. In contrast, only VMH and VMN lesions produced obesity when overeating was prevented. PFKC rats did not become obese. These experiments demonstrate that separate hypothalamic mechanisms underly the hyperphagia and obesity characteristic of VMH lesions. Furthermore, different mechanisms underly obesity and elevated parasympathetic tone following VMH lesions. Therefore, these observations support a dissociative model of the VMH obesity syndrome. / Thesis / Doctor of Philosophy (PhD)

Most of the extant humor research has focused on humor comprehension with only a few studies investigating humor appreciation as a separate construct. The purpose of this investigation was to determine the relation between humor and underlying cognitive processes. Literature on brain injured individuals has indicated that working memory, verbal and visual-spatial reasoning, cognitive flexibility, and concept formation are related to performance on comprehension tests of humor. In this study, cognitive processes underlying both verbal and nonverbal humor were investigated in a sample of healthy young adults. There is evidence that semantic and phonological humor are associated with different neural networks; therefore, both semantic and phonological humor were explored. Studies investigating physiological arousal and humor have indicated that arousal is necessary for the experience of humor. This suggests that the appreciation of humor may require the integration of cognitive and affective information, a process mediated by the ventromedial prefrontal cortex (VMPFC). Thus, a second goal of this study was to investigate the relationship between humor comprehension and appreciation and the VMPFC, by including experimental tasks that previously have been linked to VMPFC functioning. Participants included 94 undergraduate psychology students between the ages of 18 and 39 years. Participants watched film clips and listened to jokes. After the presentation of each joke and each film clip, they completed a humor comprehension/appreciation inventory developed for this study. They also completed measures assessing a range of cognitive abilities hypothesized to underlie humor perception. Hierarchical regression analyses revealed that verbal reasoning was predictive of semantic humor comprehension, indicating that verbal reasoning is a core cognitive ability for the comprehension of jokes in which the humor depends on factors other than simple word play. Cognitive measures were not predictive of phonological humor comprehension or nonverbal humor comprehension. Hierarchical regression analyses revealed that the indicators of VMPFC functioning did not correlate with either humor comprehension or humor appreciation and did not moderate the relation between humor comprehension and humor appreciation. Future research is necessary to elucidate the relationships between cognitive abilities and humor perception and to further explore the contribution of the VMPFC to humor appreciation.

Major depressive disorder (MDD) is a debilitating disorder affecting hundreds of millions of people worldwide. The etiology of the disease is unknown, and how antidepressant medications reverse depression is unclear. However, imaging and postmortem studies of MDD patients show abnormalities in several limbic areas of the brain, including the prefrontal cortex. The involvement of the ventromedial prefrontal cortex (vmPFC) in depression has been particularly intriguing, for this region demonstrates reduced metabolic activity in remission, and this reduction is unique to treatment responders. In addition, deep brain stimulation targeting the subgenual cingulate cortex in the vmPFC has been shown to be effective in treating 'treatment-resistant' patients. Furthermore, neuroanatomical studies have shown that this region projects to many downstream limbic areas implicated to play roles in MDD. I therefore hypothesized that 1) the vmPFC may be an important target of antidepressant drugs, and that 2) this region may play a role in the generation of depression-associated behaviors. To test the first hypothesis, I administered desipramine (DMI), a tricyclic antidepressant, to rats. I found that the rat vmPFC was significantly activated by DMI, whereas the dorsomedial PFC (dmPFC) was not. I also found that the drug increases neuronal activity in the nucleus accumbens, but this activation was dependent on the integrity of the vmPFC. To test the second hypothesis, I induced neuronal lesions in the rat dmPFC or vmPFC and subjected the animals to behavioral tests. I found that while lesions in both areas led to increased REM sleep, only vmPFC-lesioned animals had reduced REM latency, increased sleep fragmentation and increased forced swim test immobility. Together, these results demonstrate that the vmPFC may be an important region for both antidepressant action and the generation of depression-like behaviors.

Made available in DSpace on 2015-07-22T20:49:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-04-28. Added 1 bitstream(s) on 2015-08-11T03:26:31Z : No. of bitstreams: 1 Publico-472.pdf: 622403 bytes, checksum: 6ffd5126c843b5fda444f16935ac1337 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / O hipotálamo é uma estrutura de fundamental importância para a adaptação do organismo a situações de estresse, seja ele provocado por estímulos aversivosou por qualquer situação que altere o equilíbrio homeostático. Os núcleoshipotalâmicos mediais têm sido propostos como relacionados à modulação de respostas comportamentais de defesa e das alterações fisiológicas que a acompanham. Fazem parte do hipotálamo medial, o núcleo dorsomedial, a porção dorsal do hipotálamo ventromedial, o núcleo anterior do hipotálamo e o núcleo premamilar dorsal. A participação do sistema GABAérgico dohipotálamo medial na gênese de respostas comportamentais e fisiológicas aestímulos aversivos tem sido evidenciada por diversos estudos experimentais.Tem sido demonstrado que a administração de antagonistas GABAérgicosintra-hipotálamo medial induz comportamento de fuga, à semelhança daestimulação elétrica do núcleo. Pouco tem sido investigado com relação à modulação exercida pelo hipotálamo medial sobre respostas de defesa condicionadas e/ou evocadas por situações mais naturalísticas. O objetivo do presente trabalho foi melhor investigar o envolvimento do sistema GABAérgico e dos substratos neurais do hipotálamo medial em respostas comportamentais de defesa condicionadas e incondicionadas, a partir da utilização de um modelo animal de ansiedade, o labirinto em T elevado (LTE), que permite a medida de duas respostas comportamentais (esquiva inibitória e fuga de uma via), em um mesmo animal. Em termos clínicos, estas respostas têm sido relacionadas à ansiedade generalizada e ao transtorno do pânico, respectivamente. No presente trabalho, ratos Wistar machos (aproximadamente 280 g) foram implantados com cânulas-guia para microinjeção em dois núcleos do hipotálamo medial: o hipotálamo dorsomedial e a porção dorsal do hipotálamo ventromedial. Em um primeiro momento (experimentos 1 e 2) , os animais receberarm microinjeções do agonista GABAA muscimol (0,5 e 1,0 nmol, no experimento 1; 1,0 nmol no experimento 2) e dez minutos depois foram submetidos ao LTE. Para fins de comparação, em um terceiro experimento um grupo de animais foi tratado intra-hipotálamo dorsomedial com o anestésico local lidocaína (1 nmol; experimento 3). A fim de se evitar falsos negativos ou positivos devido ao efeito das drogas sobre a atividade locomotora, os animais foram testados em um campo aberto após os testes com o LTE. Os resultados mostraram um efeito seletivo do muscimol intra-núcleo dorsomedial sobre as respostas de fuga e esquiva. A injeção de muscimol na dose de 0,5 nmoles neste núcleo não alterou a latência sobre a esquiva inibitória dos braços abertos, nas três medidas tomadas, aumentando a latência sobre a fuga nas medidas 2 e 3. na dose de 1.0 nmol o muscimol aumentou a latência basal sobre a esquiva inibitória e a latência de fuga 3, embora tenha produzido também alteração motora verificada através da diminuição do número de cruzamentos e levantamentos no teste do campo aberto. Tal alteração motora não foi verificada na dose mais baixa. De maneira semelhante, o anestésico local lidocaína, administrado intra-núcleo dorsomedial, também não alterou nenhuma das duas medidas realizadas no LTE, esquiva e fuga, muito embora o efeito da droga não restrinja-se à inibição de corpos neuronais, já que a droga inativa também fibras de passagem. Para concluir, é possível afirmar que a inativação do hipotálamo dorsomedial por muscimol inibe seletivamente uma resposta comportamental de defesa, a fuga, que em termos clínicos tem sido relacionada ao transtorno do pânico / Previous evidence indicates that the medial hypothalamus is part of a neurobiological substrate controlling defensive behavior. In particular, it has been shown that a hypothalamic nucleus, the dorsomedial hypothalamus (DMH), is involved in the regulation of escape, a defensive behavior related to panic attacks. The role played by other hypothalamic nuclei in the organization of fear-related responses however is less clear. In this study we addressed this question by investigating the effects of the reversible inactivation of two hypothalamic nuclei, the DMH and the dorsomedial part of the ventromedial hypothalamus (VMHdm), on escape behavior generated in male Wistar rats by an ethologically relevant threatening stimulus: the exposure of rats to the open arms of the elevated T-maze. Results showed that intra-DMH administration of the GABAA receptor agonist muscimol (0.5 nmol and 1.0 nmol/0.2 μl) inhibited escape behavior, suggesting an antiaversive effect, although the higher dose also altered locomotor activity in an open field. Muscimol intra-DMH did not affect elevated T-maze inhibitory avoidance, a behavior associated with generalized anxiety disorder. On the other hand, muscimol intra-VMHdm did not alter either avoidance or escape measurements. Also, intra-DMH administration of the sodium channel blocker lidocaine (1 nmol/0.2 μl) was without effect, what is probably related to the fact the lidocaine, unlike muscimol, also inactivate fibers of passage and not only cell bodies. Taken together, our data corroborate previous evidence suggesting that the DMH is involved in the modulation of escape. Dysfunction of this regulatory mechanism may be of relevance in the genesis/maintenance of panic disorder. / FAPESP: 06/56950-3 / TEDE / BV UNIFESP: Teses e dissertações

Processing information related to the self and inferring the mental state of another person is known to involve the ventromedial prefrontal cortex (VMPFC) in both younger and older adults (Stone et al., 2008; Kelley et al., 2002; Hynes et al., 2006; Ruby et al., 2009). According to the dorsolateral prefrontal (DLPF) theory of cognitive ageing, processing of the self should not be affected by healthy adult ageing as functions related to the VMPFC remain relatively preserved compared to functions related to the DLPF cortex (MacPherson et al., 2002). Similarly, no age difference should emerge in those tasks thought to tap functions of the VMPFC. The aim of this PhD is to investigate the effect of healthy adult ageing on the ability to process information related to the self and others. A series of experiments was designed to compare the performance of younger and older adults on tasks that investigate processing and retrieval of self-related information (e.g. behaviour prediction, personality judgement, mental state inferences, self-referential). The tasks differ in the extent to which they rely on cognitive effort. The results show that ageing does not affect self-related judgements. A further series of experiments designed to investigate affective and cognitive Theory of Mind (ToM) show that the affective performance, thought to rely on VMPFC activity, is not affected by age. In contrast, the performance of older participants differs from that of younger adults on cognitive ToM task, thought to involve DLPFC brain areas. A final experiment investigated the ability to make self versus other related judgments in a confabulating patient. The results show that the ability to reflect on the self but not on others was intact. In summary, the findings demonstrate that processing self-information and making ToM inferences remains intact in older individuals and is not overtly impaired by confabulation.

Fear generalization, the generalization of fear to innocuous stimuli, is a characteristic component of pathological anxiety. For example, after returning from war, a person might begin to experience fear in response to the sound of fireworks, a stimulus typically regarded as safe. When excessive, “overgeneralization” serves as a core feature of fear and anxiety-related disorders, such as PTSD. The present collection of studies sought to investigate the neural correlates and temporal dynamics of fear generalization in humans. The first study sought to investigate the causal role of the ventromedial prefrontal cortex (vmPFC) and hippocampus in the generalization of fear. Contrary to hypotheses, individuals with focal damage to the vmPFC (N=8) or hippocampus (N=12) did not demonstrate significantly increased fear generalization relative to individuals with brain damage outside of these regions (N=16) or normal comparison participants (N=20). Potential explanations for this finding are explored. The second study sought to investigate the time course of fear generalization in humans. Participants (N = 107) completed a fear generalization task over the course of two sessions. Results indicate that fear generalization significantly increased as the duration of time between training and testing increased. This suggests that a stimulus may elicit a generalized fear response at one arbitrarily selected time point, but not another. This study establishes a novel paradigm that can be used in future work to investigate changes in the neural correlates of fear generalization over time. Fear generalization is found across an array of anxiety disorders, making it a compelling area of study. The present work highlights the dynamic nature of fear generalization in humans. Further, the present study leads to a number of questions for future research.

The neuropeptides dynorphin and cholecystokinin (CCK), and their associated pronociceptive effects were investigated in the RVM. Utilizing a nerve-injury model (SNL), RT-PCR analysis revealed increases (p<0.05) of prodynorphin mRNA, and bradyinkin, B1- and B2-receptor mRNA, post-SNL, 14-days, 2-days, and 14-days, respectively. Administration of dynorphin into the RVM produced both acute and long-lasting (>30-days) tactile hypersensitivity. Administration of the B1-antagonist, DALBK and the B2-antagonist, Hoe-140, into the RVM significantly attenuated dynorphin-induced tactile hypersensitivity. Nerve-injury induced tactile hypersensitivity was significantly reversed by RVM administration of dynorphin antiserum or the B2-antagonist, Hoe-140. These data suggest that dynorphin is up-regulated in the RVM in nerve-injury, and via the activation of bradykinin receptors in the RVM, produces abnormal pain. Like dynorphin, CCK is up-regulated in the RVM in nerve-injury, with studies suggesting that elevated levels of CCK in the RVM mediate pronociceptive activity through CCK2 receptor activation, resulting in enhanced spinal nociceptive transmission. At present, it is unknown what key neurotransmitters are mediating this RVM CCK-driven effect at the level of the spinal cord. Here, spinal cerebrospinal fluid (CSF) levels of serotonin (5-HT) and prostaglandin E2 (PGE2) were measured in the lumbar spinal cord in naïve rats following CCK administration into the RVM. Following RVM CCK microinjection, an approximate 5-fold increase in spinal (CSF) PGE2 levels was observed, as compared to baseline controls. PGE2 levels showed a progressive increase with peak levels observed at the 80-minute post-CCK injection timepoint, whereas 5-HT levels in the spinal CSF remained unchanged following CCK administration into the RVM. This release of PGE2 coincided with the timecourse for CCK-induced mechanical hypersensitivity. Administration of the CCK2-antagonist YM022 prior to CCK into the RVM, significantly attenuated (>50%) the release of PGE2 in the spinal cord. The non-selective COX-inhibitor naproxen and the 5-HT3 antagonist ondansetron, both administered intrathecally, significantly attenuated RVM CCK-induced hindpaw tactile hypersensitivity. In summary, these data suggest a bradykinin- or CCK2-receptor antagonist could be used alone or in conjunction with current therapies in the treatment of chronic pain.