Desnutrição perinatal e o controle hipotalâmico do comportamento alimentar e do metabolismo do músculo esquelético

A deficiência de nutrientes durante os períodos críticos do desenvolvimento tem sido associada com maior risco para desenvolver obesidade e diabetes Mellitus na vida adulta. Um dos mecanismos propostos refere-se à regulação do comportamento alimentar e às alterações do metabolismo energético do músculo esquelético. Recentemente, tem sido proposta a existência de uma comunicação entre o hipotálamo e o músculo esquelético a partir de sinais autonômicos que podem explicar as repercussões da desnutrição perinatal. Assim, esta revisão tem como objetivo discutir as repercussões da desnutrição perinatal sobre o comportamento alimentar e o metabolismo energético muscular e a comunicação existente entre o hipotálamo e o músculo via sinais adrenérgicos. Foram utilizadas as bases de dados MedLine/PubMed, Lilacs e Bireme, com publicações entre 2000 e 2011. Os termos de indexação utilizados foram: feeding behavior, energy metabolism, protein malnutrition, developmental plasticity, skeletal muscle e autonomic nervous system. Concluiu-se que a desnutrição perinatal pode atuar no controle hipotalâmico do comportamento alimentar e no metabolismo energético muscular, e a comunicação hipotálamo-músculo pode favorecer o desenvolvimento de obesidade e comorbidades durante o desenvolvimento.

Post-partum testosterone administration partially reverses the effects of perinatal cadmium exposure on sexual behavior in rats

This study investigated the effects of perinatal cadmium exposure on sexual behavior, organ weight, and testosterone levels in adult rats. We examined whether immediate postpartum testosterone administration is able to reverse the toxic effects of the metal. Forty pregnant Wistar rats were divided into three groups: 1) control, 2) 10 mg kg-1 cadmium chloride per day, and 3) 20 mg kg-1 cadmium chloride per day. These dams were treated on gestational days 18 and 21 and from lactation 1 to 7. Immediately after birth, half of the offspring from the experimental and control groups received 50 μl (i.p.) of 0.2% testosterone. Male sexual behavior, histological analysis and weight of organs as well as serum testosterone levels were assessed. Results showed that both cadmium doses disrupted sexual behavior in male rats, and postnatal treatment with testosterone reversed the toxic effects of 10 mg kg-1 cadmium and attenuated the effects of 20 mg kg-1 cadmium. Body weight and absolute testis, epididymis, and seminal vesicle weight were decreased by the higher cadmium dose, and testosterone supplementation did not reverse these effects. Serum testosterone levels were unaffected by both cadmium doses. No histological changes were detected in all organs analyzed. Maternal cadmium exposure effects in sexual parameters of male rat offspring were explained by the altered masculinization of the hypothalamus. We suggest that cadmium damaged cerebral sexual differentiation by its actions as an endocrine disruptor and supported by the changes discretely observed from early life during sexual development to adult life, reflected by sexual behavior. Testosterone supplementation after birth reversed some crucial parameters directly related to sexual behavior.

Chronic unpredictable mild stress alters an anxiety-related defensive response, Fos immunoreactivity and hippocampal adult neurogenesis

Previous results show that elevated T-maze (ETM) avoidance responses are facilitated by acute restraint. Escape, on the other hand, was unaltered. To examine if the magnitude of the stressor is an important factor influencing these results, we investigated the effects of unpredictable chronic mild stress (UCMS) on ETM avoidance and escape measurements. Analysis of Fos protein immunoreactivity (Fos-ir) was used to map areas activated by stress exposure in response to ETM avoidance and escape performance. Additionally, the effects of the UCMS protocol on the number of cells expressing the marker of migrating neuroblasts doublecortin (DCX) in the hippocampus were investigated. Corticosterone serum levels were also measured. Results showed that UCMS facilitates ETM avoidance, not altering escape. In unstressed animals, avoidance performance increases Fos-ir in the cingulate cortex, hippocampus (dentate gyrus) and basomedial amygdala, and escape increases Fos-ir in the dorsolateral periaqueductal gray and locus ceruleus. In stressed animals submitted to ETM avoidance, increases in Fos-ir were observed in the cingulate cortex, ventrolateral septum, hippocampus, hypothalamus, amygdala, dorsal and median raphe nuclei. In stressed animals submitted to ETM escape, increases in Fos-ir were observed in the cingulate cortex, periaqueductal gray and locus ceruleus. Also, UCMS exposure decreased the number of DCX-positive cells in the dorsal and ventral hippocampus and increased corticosterone serum levels. These data suggest that the anxiogenic effects of UCMS are related to the activation of specific neurobiological circuits that modulate anxiety and confirm that this stress protocol activates the hypothalamus-pituitary-adrenal axis and decreases hippocampal adult neurogenesis.

Maternal immune activation increases the corticosterone response to acute stress without affecting the hypothalamic monoamine content and sleep patterns in male mice offspring

Background/Aims: Early life experiences are homeostatic determinants for adult organisms. We evaluated the impact of prenatal immune activation during late gestation on the neuroimmune-endocrine function of adult offspring and its interaction with acute stress. Methods: Pregnant Swiss mice received saline or lipopolysaccharide (LPS) on gestational day 17. Adult male offspring were assigned to the control or restraint stress condition. We analyzed plasmatic corticosterone and catecholamine levels, the monoamine content in the hypothalamus, striatum and frontal cortex, and the sleep-wake cycle before and after acute restraint stress. Results and Conclusion: Offspring from LPS-treated dams had increased baseline norepinephrine levels and potentiated corticosterone secretion after the acute stressor, and no effect was observed on hypothalamic monoamine content or sleep behavior. The offspring of immune-activated dams exhibited impairments in stress-induced serotonergic and dopaminergic alterations in the striatum and frontal cortex. The data demonstrate a distinction between the plasmatic levels of corticosterone in response to acute stress and the hypothalamic monoamine content and sleep patterns. We provide new evidence regarding the influence of immune activation during late gestation on the neuroendocrine homeostasis of offspring.

Melatonin acts through MT1/MT2 receptors to activate hypothalamic Akt and suppress hepatic gluconeogenesis in rats

Melatonin can contribute to glucose homeostasis either by decreasing gluconeogenesis or by counteracting insulin resistance in distinct models of obesity. However, the precise mechanism through which melatonin controls glucose homeostasis is not completely understood. Male Wistar rats were administered an intracerebroventricular (icv) injection of melatonin and one of following: an icv injection of a phosphatidylinositol 3-kinase (PI3K) inhibitor, an icv injection of a melatonin receptor (MT) antagonist, or an intraperitoneal (ip) injection of a muscarinic receptor antagonist. Anesthetized rats were subjected to pyruvate tolerance test to estimate in vivo glucose clearance after pyruvate load and in situ liver perfusion to assess hepatic gluconeogenesis. The hypothalamus was removed to determine Akt phosphorylation. Melatonin injections in the central nervous system suppressed hepatic gluconeogenesis and increased hypothalamic Akt phosphorylation. These effects of melatonin were suppressed either by icv injections of PI3K inhibitors and MT antagonists and by ip injection of a muscarinic receptor antagonist. We conclude that melatonin activates hypothalamus-liver communication that may contribute to circadian adjustments of gluconeogenesis. These data further suggest a physiopathological relationship between the circadian disruptions in metabolism and reduced levels of melatonin found in type 2 diabetes patients.

Effects of thyroid status on NEI concentration in specific brain areas related to reproduction during the estrous cycle

We previously showed that short-term hypo- and hyperthyroidism induce changes in neuropeptide glutamic-acid-isoleucine-amide (NEI) concentrations in discrete brain areas in male rats. To investigate the possible effects of hypo- and hyperthyroidism on NEI concentrations mainly in hypothalamic areas related to reproduction and behavior, female rats were sacrificed at different days of the estrous cycle. Circulating luteinizing hormone (LH), estradiol and progesterone concentrations were measured in control, hypothyroid (hypoT, treated with PTU during 7-9 days) and hyperthyroid (hyperT, l-T4 during 4-7 days) animals. Both treatments blunted the LH surge. Hypo- and hyperthyroidism increased estradiol concentrations during proestrus afternoon (P-PM), although hypoT rats showed lower values compared to control during proestrus morning (P-AM). Progesterone levels were higher in all groups at P-PM and in the hyperT during diestrus morning (D2). NEI concentrations were lower in hypoT rats during the estrous cycle except in estrus (E) in the peduncular part of the lateral hypothalamus (PLH). They were also reduced by both treatments in the perifornical part of the lateral hypothalamus (PeFLH) during P-PM. Hypothyroidism led to higher NEI concentrations during P-PM in the organum vasculosum of the lamina terminalis and anteroventral periventricular nucleus (OVLT+AVPV). The present results indicate that NEI concentration is regulated in a complex manner by hypo- and hyperthyroidism in the different areas studied, suggesting a correlation between NEI values and the variations of gonadal steroid levels during estrous cycle. These changes could be, in part, responsible for the alterations observed in the hypothalamic-pituitary-gonadal axis in these pathologies.

Ventral premammillary nucleus as a critical sensory relay to the maternal aggression network

Maternal aggression is under the control of a wide variety of factors that prime the females for aggression or trigger the aggressive event. Maternal attacks are triggered by the perception of sensory cues from the intruder, and here we have identified a site in the hypothalamus of lactating rats that is highly responsive to the male intruder—the ventral premammillary nucleus (PMv). The PMv is heavily targeted by the medial amygdalar nucleus, and we used lesion and immediate-early gene studies to test our working hypothesis that the PMv signals the presence of a male intruder and transfers this information to the network organizing maternal aggression. PMv-lesioned dams exhibit significantly reduced maternal aggression, without affecting maternal care. The Fos analysis revealed that PMv influences the activation of hypothalamic and septal sites shown to be mobilized during maternal aggression, including the medial preoptic nucleus (likely to represent an important locus to integrate priming stimuli critical for maternal aggression), the caudal two-thirds of the hypothalamic attack area (comprising the ventrolateral part of the ventromedial hypothalamic nucleus and the adjacent tuberal region of the lateral hypothalamic area, critical for the expression of maternal aggression), and the ventral part of the anterior bed nuclei of the stria terminalis (presently discussed as being involved in controlling neuroendocrine and autonomic responses accompanying maternal aggression). These findings reveal an important role for the PMv in detecting the male intruder and how this nucleus modulates the network controlling maternal aggression.

Purinergic and glutamatergic interactions in the hypothalamic paraventricular nucleus modulate sympathetic outflow

P2X receptors are expressed on ventrolateral medulla projecting paraventricular nucleus (PVN) neurons. Here, we investigate the role of adenosine 5′-triphosphate (ATP) in modulating sympathetic nerve activity (SNA) at the level of the PVN. We used an in situ arterially perfused rat preparation to determine the effect of P2 receptor activation and the putative interaction between purinergic and glutamatergic neurotransmitter systems within the PVN on lumbar SNA (LSNA). Unilateral microinjection of ATP into the PVN induced a dose-related increase in the LSNA (1 nmol: 38 ± 6 %, 2.5 nmol: 72 ± 7 %, 5 nmol: 96 ± 13 %). This increase was significantly attenuated by blockade of P2 receptors (pyridoxalphosphate-6-azophenyl-20,40-disulphonic acid, PPADS) and glutamate receptors (kynurenic acid, KYN) or a combination of both. The increase in LSNA elicited by L-glutamate microinjection into the PVN was not affected by a previous injection of PPADS. Selective blockade of non-N-methyl-D-aspartate receptors (6-cyano-7-nitroquinoxaline-2,3-dione disodium salt, CNQX), but not N-methyl-D-aspartate receptors (NMDA) receptors (DL-2-amino-5-phosphonopentanoic acid, AP5), attenuated the ATP-induced sympathoexcitatory effects at the PVN level. Taken together, our data show that purinergic neurotransmission within the PVN is involved in the control of SNA via P2 receptor activation. Moreover, we show an interaction between P2 receptors and non-NMDA glutamate receptors in the PVN suggesting that these functional interactions might be important in the regulation of sympathetic outflow

The cardiovascular actions of fractalkine/CX3CL1 in the hypothalamic paraventricular nucleus are attenuated in rats with heart failure

The paraventricular nucleus (PVN) of the hypothalamus plays an important role in the regulation of sympathetic nerve activity, which is significantly elevated in chronic heart failure (CHF). Fractalkine (FKN) and its cognate receptor, CX3CR1, are constitutively expressed in the central nervous system, but their role and physiological significance are not well known. The aims of the present study were to determine whether FKN plays a cardiovascular role within the PVN and to investigate how the actions of FKN might be altered in CHF. We show that both FKN and CX3CR1 are expressed on neurons in the PVN of rats, suggesting that they may have a physiological function in this brain nucleus. Unilateral microinjection of FKN directly into the PVN of anaesthetized rats elicited a significant dose-related decrease in blood pressure (1.0 nmol, -5 ± 3 mmHg; 2.5 nmol, -13 ± 2 mmHg; 5.0 nmol, -22 ± 3 mmHg; and 7.5 nmol, -32 ± 3 mmHg) and a concomitant increase in heart rate (1.0 nmol, 6 ± 3 beats min(-1); 2.5 nmol, 11 ± 3 beats min(-1); 5 nmol, 18 ± 4 beats min(-1); and 7.5 nmol, 27 ± 5 beats min(-1)) compared with control saline microinjections. In order to determine whether FKN signalling is altered in rats with CHF, we first performed quantitative RT-PCR and Western blot analysis and followed these experiments with functional studies in rats with CHF and sham-operated control rats. We found a significant increase in CX3CR1 mRNA and protein expression, as determined by quantitative RT-PCR and Western blot analysis, respectively, in the PVN of rats with CHF compared with sham-operated control rats. We also found that the blood pressure effects of FKN (2.5 nmol in 50 nl) were significantly attenuated in rats with CHF (change in mean arterial pressure, -6 ± 3 mmHg) compared with sham-operated control rats (change in mean arterial pressure, -16 ± 6 mmHg). These data suggest that FKN and its receptor, CX3CR1, modulate cardiovascular function at the level of the PVN and that the actions of FKN within this nucleus are altered in heart failure

Leptin receptors in human skeletal muscle

[EN] Human skeletal muscle expresses leptin receptor mRNA; however, it remains unknown whether leptin receptors (OB-R) are also expressed at the protein level. Fourteen healthy men (age = 33.1 +/- 2.0 yr, height = 175.9 +/- 1.7 cm, body mass = 81.2 +/- 3.8 kg, body fat = 22.5 +/- 1.9%; means +/- SE) participated in this investigation. The expression of OB-R protein was determined in skeletal muscle, subcutaneous adipose tissue, and hypothalamus using a polyclonal rabbit anti-human leptin receptor. Three bands with a molecular mass close to 170, 128, and 98 kDa were identified by Western blot with the anti-OB-R antibody. All three bands were identified in skeletal muscle: the 98-kDa and 170-kDa bands were detected in hypothalamus, and the 98-kDa and 128-kDa bands were detected in thigh subcutaneous adipose tissue. The 128-kDa isoform was not detected in four subjects, whereas in the rest its occurrence was fully explained by the presence of intermuscular adipose tissue, as demonstrated using an anti-perilipin A antibody. No relationship was observed between the basal concentration of leptin in serum and the 170-kDa band density. In conclusion, a long isoform of the leptin receptor with a molecular mass close to 170 kDa is expressed at the protein level in human skeletal muscle. The amount of 170-kDa protein appears to be independent of the basal concentration of leptin in serum.

Influenza dell'Omeostasi Idrico-Elettrolitica sul Ciclo Veglia-Sonno

During the wake sleep (W-S) cycle in mammals, the alternation of the different states, wake, NREM sleep (NREMS) and REM sleep (REMS), is associated not only with electroencephalographic or behavioural changes, but also with modifications in the physiological regulations of the organism. The most evident change is the existence of a suspension of the somatic and autonomic thermoregulatory responses during REMS. Since thermoregulation is prevalently controlled by the Preoptic Area-Anterior Hypothalamus (PO-AH), its suspension during REM sleep has been taken as a sign of an impairment of the hypothalamic integrative activity that could explain the modifications in physiological regulation observed in this sleep stage. The recent finding from our laboratory that the secretion of the antidiuretic hormone arginine-vasopressin (AVP) in response to a central osmotic stimulation is quantitatively the same throughout the different stages of the W-S cycle, has shown that hypothalamic osmoregulation is not suspended during REMS. In order to clarify the extent of the hypothalamic involvement in the regulation of the W-S cycle, we have studied the effects of three days of water deprivation and of two days of recovery during which animals were allowed a free access to water, on the architecture of the W-S cycle. The condition of water deprivation represents a severe challenge involving neuroendocrine and autonomic hypothalamic regulations. In contradiction with thermoregulatory studies, in which it has been clearly demonstrated that a thermal challenge selectively reduces REMS occurrence, the results of this study show that REMS occurrence is mildly reduced only in the third day of water deprivation. The most striking effects produced by water deprivation appear to concern NREMS, which shows a selective and significant reduction in its slow EEG activity (delta-power) but not in its duration. The recovery period is mainly characterized by a disruption of the normal circadian rhythm of REMS occurrence and by a rebound of the delta power in NREMS. Thus, an autonomic challenge different from those related to thermoregulation and an endocrine challenge as the continuous secretion of AVP show to exert different effects on the stages of the wake-sleep cycle. Also, this study demonstrates that the impairment of the hypothalamic integrative activity thought to characterize the occurrence of REMS only involves thermoregulatory structures.

Sleep-related changes in blood pressure in hypocretin-deficient narcoleptic mice

Objectives. Blood pressure (BP) physiologically has higher and lower values during the active and rest period, respectively. Subjects failing to show the appropriate BP decrease (10-20%) on passing form diurnal activity to nocturnal rest and sleep have increased risk of target organ damage at the cardiac, vascular and cerebrovascular levels. Hypocretin (HCRT) releasing neurons, mainly located in the lateral hypothalamus, project widely to the central nervous system. Thus HCRT neurons are involved in several autonomic functions, including BP regulation. HCRT neurons also play a key role in wake-sleep cycle regulation, the lack of which becomes evident in HCRT-deficient narcoleptic patients. I investigated whether chronic lack of HCRT signaling alters BP during sleep in mouse models of narcolepsy. Methods. The main study was performed on HCRT-ataxin3 transgenic mice (TG) with selective post-natal ablation of HCRT neurons, HCRT gene knockout mice (KO) with preserved HCRT neurons, and Wild-Type control mice (WT) with identical genetic background. Experiments where replicated on TG and WT mice with hybrid genetic background (hTG and hWT, respectively). Mice were implanted with a telemetric pressure transducer (TA11PA-C10, DSI) and electrodes for discriminating wakefulness (W), rapid-eye-movement sleep (REMS) and non-REMS (NREMS). Signals were recorded for 3 days. Mean BP values were computed in each wake-sleep state and analyzed by ANOVA and t-test with significance at p<0.05. Results. The decrease in BP between either NREMS or REMS and W was significantly blunted in TG and KO with respect to WT as well as in hTG with respect to hWT. Conclusions. Independently from the genetic background, chronic HCRT deficiency leads to a decreased BP difference between W and sleep potentially adverse in narcoleptic subjects. These data suggest that HCRT play an important role in the sleep-dependent cardiovascular control.

Effects of ambient temperature on cardiovascular regulation during sleep in hypocretin-deficient narcoleptic mice

Hypocretin 1 and 2 (HCRT, also called Orexin A and B) are neuropeptides released by neurons in the lateral hypothalamus. HCRT neurons widely project to the entire neuroaxis. HCRT neurons have been reported to participate in various hypothalamic physiological processes including cardiovascular functions, wake-sleep cycle, and they may also influence metabolic rate and the regulation of body temperature. HCRT neurons are lost in narcolepsy, a rare neurological disorder, characterized by excessive daytime sleepiness, cataplexy, sleep fragmentation and occurrence of sleep-onset rapid-eye-movement episodes. We investigated whether HCRT neurons mediate the sleep-dependent cardiovascular adaptations to changes in ambient temperature (Ta). HCRT-ataxin3 transgenic mice with genetic ablation of HCRT neurons (n = 11) and wild-type controls (n = 12) were instrumented with electrodes for sleep scoring and a telemetric blood pressure (BP) transducer (DSI, Inc.). Simultaneous sleep and BP recordings were performed on mice undisturbed and freely-behaving at 20 °C, 25 °C, and 30 °C for 48 hours at each Ta. Analysis of variance of BP indicated a significance of the main effects of wake-sleep state and Ta, their interaction effect, and the wake-sleep state x mouse strain interaction effect. BP increased with decreasing Ta. This effect of Ta on BP was significantly lower in rapid-eye-movement sleep (REMS) than either in non-rapid-eye-movement sleep (NREMS) or wakefulness regardless of the mouse strain. BP was higher in wakefulness than either in NREMS or REMS. This effect of sleep on BP was significantly reduced in mice lacking HCRT neurons at each Ta, particularly during REMS. These data suggest that HCRT neurons play a critical role in mediating the effects of sleep but not those of Ta on BP in mice. HCRT neurons may thus be part of the central neural pathways which mediate the phenomenon of blood pressure dipping on passing from wakefulness to sleep.

Effetti sulle funzioni autonomiche e sugli stati di veglia e di sonno della manipolazione centrale farmacologica del sistema ipocretinergico nel ratto

Obiettivo della tesi è stato quello di studiare il ruolo svolto dall’ipotalamo laterale (LH) nella regolazione dei processi di integrazione dell’attività autonomica e termoregolatoria con quella degli stati di veglia e sonno. A questo scopo, l’attività dell’LH è stata inibita per 6 ore (Esperimento A) mediante microiniezioni locali dell’agonista GABAA muscimolo nel ratto libero di muoversi, nel quale sono stati monitorati in continuo l’elelttroencefalogramma, l’elettromiogramma nucale, la pressione arteriosa (PA) e la temperatura ipotalamica (Thy) e cutanea. Gli animali sono stati studiati a temperatura ambientale (Ta) di 24°C e 10°C. I risultati hanno mostrato che l’inibizione acuta dell’LH riduce l’attività di veglia e sopprime la comparsa del sonno REM. Ciò avviene attraverso l’induzione di uno stato di sonno NREM caratterizzato da ipersincronizzazione corticale, con scomparsa degli stati transizionali al sonno REM. Quando l’animale è esposto a bassa Ta, tali alterazioni si associano a un ampio calo della Thy, che viene compensato da meccanismi vicarianti solo dopo un paio d’ore dall’iniezione. Sulla base di tali risultati, si è proceduto ad un ulteriore studio (Esperimento B) volto ad indagare il ruolo del neuropeptide ipocretina (prodotto in modo esclusivo a livello dell’LH) nei processi termoregolatori, mediante microiniezioni del medesimo nel bulbo rostrale ventromediale (RVMM), stazione cruciale della rete nervosa preposta all’attivazione dei processi termogenetici. La somministrazione di ipocretina è stata in grado di attivare la termogenesi e di potenziare la comparsa della veglia, con concomitante lieve incremento della PA e della frequenza cardiaca, quando effettuata alle Ta di 24°C o di 10°C, ma non alla Ta di 32°C. In conclusione, i risultati indicano che l’LH svolge un ruolo cruciale nella promozione degli stati di veglia e di sonno REM e, per tramite dell’ipocretina, interviene in modo coplesso a livello del RVMM nella regolazione dei processi di coordinamento dell'attività di veglia con quella termoregolatoria.

Untersuchungen zur Lokalisation und Expression des respiratorischen Proteins Neuroglobin bei Säugetieren

In der vorliegenden Arbeit wurde Neuroglobin (Ngb), ein evolutiv altes und in Metazoen konserviertes respiratorisches Protein, funktionell untersucht. Mittels des induzierbaren Tet on / Tet off Systems wurde Ngb ektopisch in der murinen Leber und im Gehirn überexprimiert. Die Transkriptome von Leber und Gehirnregionen Ngb-transgener Mäuse wurden mittels Microarrays und RNA-Seq im Vergleich zum Wildtyp analysiert, um Auswirkungen der Ngb-Überexpression zu ermitteln. Die Transkriptom-Analyse in Leber und Gehirn zeigte eine nur geringe Anzahl differenziell regulierter Gene und Stoffwechselwege nach Ngb-Überexpression. Ngb transgene Mäuse wurden CCl4-induziertem ROS-Stress ausgesetzt und die Leberfunktion untersucht. Zudem wurden primäre Hepatozyten-Kulturen etabliert und in diesen in vitro die extrinsische Apoptose induziert. Die Stressversuche zeigten: (i) Die Ngb-Überexpression hat keine protektive Wirkung in der Leber in vivo. (ii) In Leberzellen in vitro hingegen verminderte eine Ngb-Überexpression effizient die Aktivierung der apoptotischen Kaskade. Eine protektive Wirkung von Ngb ist vermutlich von betrachtetem Gewebe und dem verwendeten Stressor abhängig und keine generelle, selektierte Funktion des Proteins.rnWeiterhin wurde eine Ngb-KnockOut-Mauslinie mit einem LacZ-KnockIn-Genotyp etabliert. Hierbei zeigten die KO-Mäuse keinen offensichtlichen Phänotyp in ihrer Entwicklung, Fortpflanzung und Retina-Funktion. Unter Verwendung des LacZ-Knockin-Konstrukts konnten kontrovers diskutierte Ngb-Expressionsorte im adulten Mausgehirn (Hippocampus, Cortex und Cerebellum) sowie in Testes experimentell bestätigt werden. Parallel wurden öffentlich verfügbare RNA-Seq Datensätze ausgewertet, um die regionale Ngb-Expression systematisch ohne Antikörper-assoziierte Spezifitätsprobleme zu charakterisieren. Eine basale Ngb-Expression (RPKM: ~1-5) wurde im Hippocampus, Cortex und Cerebellum, sowie in Retina und Testes ermittelt. Eine 20-40fach höhere, starke Expression (RPKM: ~160) wurde im Hypothalamus bzw. im Hirnstamm nachgewiesen. Die „digitale“ Expressionsuntersuchung wurde mittels qRT-PCR und Western Blot bestätigt. Dieses Expressionsprofil von Ngb in der Maus weist auf eine besondere funktionelle Bedeutung von Ngb im Hypothalamus hin. Eine Funktion von Ngb in der Sauerstoffversorgung der Retina und eine generelle Funktion von Ngb in der Protektion von Neuronen sind mit dem beobachteten Expressionsspektrum weniger gut vereinbar.