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Effects of hypocretin (orexin) neuronal loss on sleep and extracellular adenosine levels in the rat basal forebrain.
Murillo-Rodriguez E, Liu M, Blanco-Centurion C, Shiromani PJ (2008) Effects of hypocretin (orexin) neuronal loss on sleep and extracellular adenosine levels in the rat basal forebrain. Eur J Neurosci 28(6):1191-1198. doi: 10.1111/j.1460-9568.2008.06424.x
Summary: Adenosine levels in the basal forebrain are thought to regulate the waxing and waning of sleep drive. Rats received bilateral 100-ng injections of orexin-SAP (Cat. #IT-20) into the lateral hypothalamus – resulting in a 94% loss of orexin-containing neurons. Lesioned animals displayed several changes in sleep characteristics, but no increase of adenosine levels after sleep deprivation. The results indicate that sleep changes due to orexin-SAP lesioning occur independently of adenosine levels.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Effects of saporin-induced lesions of three arousal populations on daily levels of sleep and wake.
Blanco-Centurion C, Gerashchenko D, Shiromani PJ (2007) Effects of saporin-induced lesions of three arousal populations on daily levels of sleep and wake. J Neurosci 27:14041-14048. doi: 10.1523/JNEUROSCI.3217-07.2007
Summary: Orexin neurons in the basal forebrain, tuberomammillary nucleus (TMN), and locus ceruleus (LC) are thought to regulate arousal. Rats were injected with two or three of the following targeted conjugates: anti-DBH-SAP (Cat. #IT-03), 0.25 µl bilateral injections of 1 µg/µl into the LC; orexin-SAP (Cat. #IT-20), 0.25 µl injection of 0.25 µg/µl into the TMN; 192-IgG-SAP (Cat. #IT-01), 3 µl injection of 2 µg/µl into the lateral ventricle. Small differences were observed in sleep architecture, but the data do not support the traditional hypothesis that these three areas of the brain are essential links in the control of wake levels.
Related Products: Anti-DBH-SAP (Cat. #IT-03), Orexin-B-SAP (Cat. #IT-20), 192-IgG-SAP (Cat. #IT-01)
Food-elicited increases in cortical acetylcholine release require orexin transmission.
Frederick-Duus D, Guyton MF, Fadel J (2007) Food-elicited increases in cortical acetylcholine release require orexin transmission. Neuroscience 149:499-507. doi: 10.1016/j.neuroscience.2007.07.061
Summary: In these experiments the authors examine the hypothesis that orexin fibers from the hypothalamus are necessary for basal forebrain cholinergic system (BFCS) activation in a food restriction model. Rats received 200 ng of orexin-SAP (Cat. #IT-20) into the lateral hypothalamus/perifornical area. Lesioned animals that were also deprived of food displayed increased feeding latency when presented with food. These and other data indicate orexin in the BFCS is involved in attention to stimuli associated with homeostatic challenges.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Transplant of hypocretin neurons into the lateral hypothalamus of a narcolepsy rat model
Millan-Aldaco D, Arias-Carrion O, Palomero-Rivero M, Drucker-Colin R, Murillo-Rodriguez E (2007) Transplant of hypocretin neurons into the lateral hypothalamus of a narcolepsy rat model. Neuroscience 2007 Abstracts 779.2/E4. Society for Neuroscience, San Diego, CA.
Summary: Narcolepsy, a disabling neurological disorder, is characterized by excessive daytime sleepiness, sleeps attacks, sleep fragmentation, and cataplexy. This sleep disorder has been linked to a loss of neurons into the lateral hypothalamus (LH) containing the neuropeptide hypocretin (HCRT). Our group has developed an experimental model in rats that mimics several aberrant behaviours observed in human narcolepsy. The bilateral administration of the neurotoxin hypocretin-2-saporin (HCRT2-SAP) into the LH of rats destroys most of the HCRT neurons (~90%) leading to develop narcolepsy as evaluated using EEG/EMG means. In order to replace the HCRT lost neurons by the local injection of the HCRT2-SAP, a suspension of cells from the hypothalamus obtained from rat pups (3-5 days old) were processed for grafting and stained with GFP. This cell suspension was injected into the LH of lesioned rats and they were sacrificed 21 days post-transplant. The brain was cut and sections containing LH were processed for HCRT immunohistochemistry as well as for the presence of HCRT-immunoflorescence neurons. We were able to differentiate the HCRT transplanted neurons into the LH of lesioned rats. Importantly, they were present at the target area 21 days after implant. These somata were similar in size and appearance to adult rat HCRT-immunoreactive neurons. Our results are very promising since the present study indicates that HCRT neurons obtained from rat pups can be grafted into a host brain and graft survived during 21 days. This experimental approach definitely addresses the possibility to replace HCRT neurons in narcolepsy in order to reverse this disease.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Early activation of the tuberomammillary nucleus is a common factor in appetitive behaviors in rats
Contreras M, Carrasco M, Riveros M, Quispe M, Valdes J, Torrealba F (2007) Early activation of the tuberomammillary nucleus is a common factor in appetitive behaviors in rats. Neuroscience 2007 Abstracts 842.19/WW11. Society for Neuroscience, San Diego, CA.
Summary: Histamine neurons of the tuberomammillary nucleus show an earlier activation during the appetitive phase of feeding, compared to the other arousal system nuclei. To test if in different appetitive behaviors also these histaminergic neurons become active first, we studied changes in Fos-ir in arousal nuclei during sexual, drinking and drug-seeking behavior. Male rats were exposed to sexually receptive or to non-receptive female rats, allowing sensory but not sexual contact. Receptive females elicited increased sniffing time which positively correlated with Fos-ir in the dorsal raphe, laterodorsal tegmental nucleus, orexin hypothalamic neurons and tuberomammillary nucleus. Non receptive females induced less sniffing and no increased Fos-ir. Other male rats were deprived of water for 48 h and presented with an empty water bottle to induce appetitive behavior. The presentation of an empty water bottle to thirsty rats induced increased approaches to the bottle while they tried to drink. While water deprivation per se increased Fos-ir in the dorsal raphe and the locus coeruleus, the presentation of the bottle increased Fos-ir in the tuberomammillary nucleus and induced a further Fos-ir increase in the locus coeruleus. Other male rats were conditioned to amphetamine (1.5 mg/Kg i.p.) using a place preference task. Conditioned rats, but not rats injected with saline instead of amphetamine, showed a significant preference for amphetamine-paired room and increase in the number of Fos-ir in the tuberomammillary nucleus, orexin hypothalamic neurons and locus coeruleus. To evaluate the importance of the histaminergic neurons in the appetitive phase of these motivated behavior, we lesioned tuberomammillary nucleus using saporin conjugated to the hypocretin 2. The histaminergic neurons lesion blunted the appetitive phase in all motivated behaviors studied, without affecting general motor capacities. Taken together our results indicate that the histaminergic neurons become active at the onset of different motivated behaviors and they are key in the arousal that is essential in motivation. Other arousal nuclei may participate depending on the particular behavior.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Genetic dissection of neural circuitry underlying REM sleep behavior disorder (RBD)
Wood DA, Patterson N, Fuller P, Sherman D, Saper C, Lu J (2007) Genetic dissection of neural circuitry underlying REM sleep behavior disorder (RBD). Neuroscience 2007 Abstracts 736.28/VV11. Society for Neuroscience, San Diego, CA.
Summary: REM sleep behavior disorder (RBD), a parasomnia typically manifested as dream enactment behavior, may represent an early pathophysiologic manifestation of Lewy body diseases (LBD), such as Parkinson disease and dementia with Lewy bodies. Preclinical investigation of possible underlying neural mechanisms of RBD suggests that a set of glutamatergic neurons located in the sublaterodorsal nucleus (SLD), which project to GABA/glycine interneurons in the ventral horn are responsible for atonia during REM sleep (Lu et al. 2006, A putative flip-flop switch for control of REM sleep, Nature 441, 589-94). Based upon these findings, we hypothesize that a loss of glutamate from these neurons in the SLD produces REM sleep without atonia, an animal equivalent of RBD. To assess this question, we selectively eliminated glutamate release from SLD by injecting adeno-associated virus-Cre recombinase (AAV-Cre) into the SLD of mice with lox P sites flanking exon 2 of the vesicular glutamate transporter 2 (VGLUT2) gene. In addition, we examined the role of the ventromedial medulla (VMM) in REM atonia by injecting orexin-saporin in rats and AAV-Cre into flox-VGAT (vesicular GABA/glycine transporter) and flox-VGLUT2 mice. Consistent with our hypothesis, these data show that loss of the VGLUT2 gene in the SLD produces REM sleep without atonia (walking, running and myoclonic jerking) without alteration of total amount of REM sleep. Furthermore, loss of the VGLUT2 but not the VGAT gene in the intermediate VMM results in myoclonic jerking against the background of tonic atonia during REM sleep. Based upon these observations, we propose that suppression of muscle activity during REM sleep is controlled by the activation of excitatory glutamatergic projections from the SLD (with collaterals targeting the intermediate VMM) and from the intermediate VMM, which terminate at inhibitory interneurons in the spinal cord. Collectively, this work provides novel insight into the control of muscle tone during REM sleep, which may have implications for our understanding of neurological conditions that precede the onset of neurodegenerative disease.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Hypocretin/orexin neuronal loss increases adult neurogenesis
Arias-Carrion O, Hernandez-Martinez H, Drucker-Colin R (2007) Hypocretin/orexin neuronal loss increases adult neurogenesis. Neuroscience 2007 Abstracts 456.14/C7. Society for Neuroscience, San Diego, CA.
Summary: Adult neurogenesis in the subventricular zone (SVZ) is subjected to physiological regulation and can be modified by brain injuries. The sleep disorder narcolepsy may now be considered a neurodegenerative disease, as there is a massive reduction in the number of neurons containing the neuropeptide, hypocretin (HCRT). In the present study, we investigate the relationship between hypocretin neuronal loss and adult neurogenesis. The neurotoxin, hypocretin-2-saporin (HCRT2-SAP), was administered bilaterally to the lateral hypothalamus (LH) to lesion HCRT neurons. Five weeks after HCRT2-SAP administration a loss of HCRT-ir neurons into LH was produced. In normal animals, a high density of HCRT-ir fibers was found in the septum and was poor in the corpus callosum and striatum. These densities decreased in lesioned animals. To label dividing cells, we used 5-bromo-2′-deoxyuridine (BrdU). BrdU was injected twice daily during days 10-14 after lesion, saline or control procedure. Animals were killed at 3 weeks after the last BrdU injection. Experimental depletion of HCRT in rats increases precursor cell proliferation in the SVZ and subependimal layer of 3rd ventricle. However, we don’t find BrdU/HCRT double-labeled cells in the subependimal zone or LH. These observations suggest that the HCRT is a negative factor in adult neurogenesis.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Neurotoxic lesions centered on the perifornical hypothalamus abolish the cardiovascular and behavioral responses of conditioned fear to context but not of restraint.
Furlong T, Carrive P (2007) Neurotoxic lesions centered on the perifornical hypothalamus abolish the cardiovascular and behavioral responses of conditioned fear to context but not of restraint. Brain Res 1128(1):107-119. doi: 10.1016/j.brainres.2006.10.058
Summary: This work examined the role of orexin-containing neurons in the perifornical hypothalamus (PeF) during stress response. Orexin-SAP (Cat. #IT-20) or the control conjugate blank-SAP (Cat. #IT-21) was injected into the PeF of pre-conditioned rats. Tests measuring restraint and conditioned fear to context were then performed on the lesioned animals. While the lesioning was not specific enough to connect results to orexin-containing neurons, the data indicate that the PeF is critical for some forms of stress, but not others.
Related Products: Orexin-B-SAP (Cat. #IT-20), Blank-SAP (Cat. #IT-21)
Transplant of hypocretin neurons into the lateral hypothalamus of rats with lesions of the hypocretin neurons
Hernandez-Martinez H, Arias-Carrion O, Drucker-Colin R, Murillo-Rodriguez E (2006) Transplant of hypocretin neurons into the lateral hypothalamus of rats with lesions of the hypocretin neurons. Neuroscience 2006 Abstracts 719.2. Society for Neuroscience, Atlanta, GA.
Summary: Narcolepsy, a disabling neurological disorder is characterized by excessive daytime sleepiness, sleeps attacks, sleep fragmentation, and cataplexy. This sleep disorder has been linked to a loss of neurons containing the neuropeptide hypocretin (HCRT). Our group has developed an experimental model to induce narcolepsy in rats. The bilateral administration of the neurotoxin hypocretin-2-saporin (HCRT2-SAP) into the lateral hypothalamus (LH) of rats destroys the HCRT neurons. Therefore, the loss of HCRT neurons leads to developing narcolepsy. In order to replace the HCRT lost neurons by HCRT2-SAP, a suspension of cells from the posterior hypothalamus of 3-5 days old rat pups were stained with GFP and injected into the LH of lesioned rats. Animals were sacrificed 21 days after transplant, and cryostat-cut coronal sections of the LH sections were examined for presence of HCRT-immunofluorescence neurons. Preliminary data shows that HCRT transplanted neurons into the LH of lesioned rats were present at the target area 21 days after implant. These somata were similar in size and appearance to adult rat HCRT-immunoreactive neurons. Our results indicate that HCRT neurons obtained from rat pups can be grafted into a host brain and graft survives during 21 days. Importantly, our study addresses the possibility to replace HCRT neurons in narcolepsy in order to reverse this disease.
Related Products: Orexin-B-SAP (Cat. #IT-20)
Orexin transmission is required for food-related increases in cortical acetylcholine release
Frederick-Duus D, Fadel J (2006) Orexin transmission is required for food-related increases in cortical acetylcholine release. Neuroscience 2006 Abstracts 369.23. Society for Neuroscience, Atlanta, GA.
Summary: The hypothalamic orexin/hypocretin neuropeptides (OxA and OxB) are crucial modulators of state-dependent behavior including the regulation of arousal in response to homeostatic challenges. Orexins provide a moderately dense innervation of cholinergic portions of the basal forebrain, including the ventral pallidum/substantia innominata and nucleus basalis magnocellularis. OxA administration in this area also produces robust increases in cortical acetylcholine (ACh) release. Here, we used in vivo microdialysis to test the hypothesis that orexin transmission is required for the increase in cortical ACh release resulting from presentation of stimuli related to palatable food. Rats were mildly food-deprived and trained to associate sudden darkness in the testing room with presentation of sweetened cereal. Stimulated cortical ACh release in these animals was blocked by orexin B-saporin (OxB-SAP) lesions of the perifornical hypothalamus at doses that produced 75-80% loss of orexin neurons, but minimal loss of other neuronal phenotypes in this area. In intact animals, pretreatment with the orexin 1 receptor (Ox1R) antagonist SB334867 similarly abolished food cue-elicited increases in cortical ACh release, indicating the specific involvement of OxA in this phenomenon. Neither OxB-SAP nor SB334867 reduced affected basal ACh release. Finally, in old rats (28-30 months), double-label immunohistochemistry revealed a reduction in orexin-immunoreactive fibers near cholinergic somata and dendrites in the basal forebrain regions, consistent with the deficits in stimulated ACh release seen with old animals in this paradigm. Collectively, these data suggest that phasic orexin activation of the basal forebrain cholinergic system may bias attentional resources toward stimuli related to underlying homeostatic challenges, thus coordinating the processing of interoceptive and exteroceptive cues. Age-related deficits in these capacities may have an orexinergic basis.
Related Products: Orexin-B-SAP (Cat. #IT-20)