Oliver Selbach

Histamine in the Nervous System

Histamine is a transmitter in the nervous system and a signaling molecule in the gut, the skin, and the immune system. Histaminergic neurons in mammalian brain are located exclusively in the tuberomamillary nucleus of the posterior hypothalamus and send their axons all over the central nervous system. Active solely during waking, they maintain wakefulness and attention. Three of the four known histamine receptors and binding to glutamate NMDA receptors serve multiple functions in the brain, particularly control of excitability and plasticity. H1 and H2 receptor-mediated actions are mostly excitatory; H3 receptors act as inhibitory auto- and heteroreceptors. Mutual interactions with other transmitter systems form a network that links basic homeostatic and higher brain functions, including sleep-wake regulation, circadian and feeding rhythms, immunity, learning, and memory in health and disease.

Functions of neuronal adenosine receptors

Endogenous adenosine in nervous tissue, a central link between energy metabolism and neuronal activity, varies according to behavioral state and (patho)physiological conditions; it may be the major sleep propensity substance. The functional consequences of activation of the four known adenosine receptors, A1, A2A, A2B and A3, are considered here. The mechanisms and electrophysiological actions, mainly those of the A1-receptor, have been extensively studied using in vitro brain-slice preparations. A1-receptor activation inhibits many neurons postsynaptically by inducing or modulating ionic currents and presynaptically by reducing transmitter release. A1-receptors are almost ubiquitous in the brain and affect various K+ (Ileak, IAHP), mixed cationic (Ih), or Ca2+ currents, through activation of Gi/o-proteins (coupled to ion channels, adenylyl cyclase or phospholipases). A2A-receptors are much more localized, their functional role in the striatum is only just emerging. A2B- and A3-receptors may be affected in pathophysiological events, their function is not yet clear. The cAMP-PKA signal cascade plays a central role in the regulation of both neural activity and energy metabolism. Under conditions of increased demand and decreased availability of energy (such as hypoxia, hypoglycemia and/or excessive neuronal activity), adenosine provides a powerful protective feedback mechanism. Interaction with adenosine metabolism is a promising target for therapeutic intervention in neurological and psychiatric diseases such as epilepsy, sleep, movement (parkinsonism or Huntingtons disease) or psychiatric disorders (Alzheimers disease, depression, schizophrenia or addiction).

The G‐protein coupled bile salt receptor TGR5 is expressed in liver sinusoidal endothelial cells

Sinusoidal endothelial cells (SEC) constitute a permeable barrier between hepatocytes and blood. SEC are exposed to high concentrations of bile salts from the enterohepatic circulation. Whether SEC are responsive to bile salts is unknown. TGR5, a G‐protein–coupled bile acid receptor, which triggers cAMP formation, has been discovered recently in macrophages. In this study, rat TGR5 was cloned and antibodies directed against the C‐terminus of rat TGR5 were developed, which detected TGR5 as a glycoprotein in transfected HepG2‐cells. Apart from Kupffer cells, TGR5 was detected in SEC of rat liver. SEC expressed TGR5 over the entire acinus, whereas endothelial cells of the portal or central veins were not immunoreactive toward TGR5 antibodies. In isolated SEC, TGR5 mRNA and protein were detected by reverse transcription (RT) PCR, immunofluorescence microscopy, and Western blot analysis. Bile salts increased cAMP in isolated SEC and induced mRNA expression of endothelial NO synthase (eNOS), a known cAMP‐dependent gene. In addition, bile acids activated eNOS by phosphorylation of eNOS at amino acid position 1177. In line with eNOS activation, bile acids induced NO production in liver slices. This is the first report on the expression of TGR5 in SEC. Conclusion: The data suggest that SEC are directly responsive toward specific bile salts. Regulation of eNOS in SEC by TGR5 connects bile salts with hepatic hemodynamics. This is of particular importance in cholestatic livers when bile salt concentrations are increased. (HEPATOLOGY 2007;45:695–704.)

Hypoosmotic swelling and ammonia increase oxidative stress by NADPH oxidase in cultured astrocytes and vital brain slices.

The role of NADPH oxidase (NOX) and the regulatory subunit p47phox for hypoosmotic ROS generation was studied in cultured rat astrocytes and brain slices of wilde type and p47phox knock‐out mice. Cultured rat astrocytes express mRNAs encoding for the regulatory subunit p47phox, NOX1, 2, and 4, and the dual oxidases (DUOX)1 and 2, but not NOX3. Hypoosmotic (205 mosmol/L) swelling of cultured astrocytes induced a rapid generation of ROS that was accompanied by serine phosphorylation of p47phox and prevented by the NADPH oxidase inhibitor apocynin. Apocynin also impaired the hypoosmotic tyrosine phosphorylation of Src. Both, hypoosmotic ROS generation and p47phox serine phosphorylation were sensitive to the acidic sphingomyelinase inhibitors AY9944 and desipramine, the protein kinase C (PKC)ζ‐inhibitory pseudosubstrate peptide, the NMDA receptor antagonist MK‐801 and the intracellular Ca2+ chelator BAPTA‐AM. Also hypoosmotic exposure of wilde type mouse cortical brain slices increased ROS generation, which was allocated in part to the astrocytes and which was absent in presence of apocynin and in cortical brain slices from p47phox knock‐out mice. Also ammonia induced a rapid ROS production in cultured astrocytes and brain slices, which was sensitive to apocynin. The data suggest that astrocyte swelling triggers a p47phox‐dependent NADPH oxidase‐catalyzed ROS production. The findings further support a close interrelation between osmotic and oxidative stress in astrocytes, which may be relevant to different brain pathologies including hepatic encephalopathy.

Temporal dynamics of mouse hippocampal clock gene expression support memory processing

Hippocampal plasticity and mnemonic processing exhibit a striking time‐of‐day dependence and likely implicate a temporally structured replay of memory traces. Molecular mechanisms fulfilling the requirements of sensing time and capturing time‐related information are coded in dynamics of so‐called clock genes and their protein products, first discovered and described in the hypothalamic suprachiasmatic nucleus. Using real‐time PCR and immunohistochemical analyses, we show that in wildtype mice core clock components (mPer1/PER1, mPer2/PER2, mCry1/CRY1, mCry2/CRY2, mClock/CLOCK, mBmal1/BMAL1) are expressed in neurons of all subregions of the hippocampus in a time‐locked fashion over a 24‐h (diurnal) day/night cycle. Temporal profiling of these transcriptional regulators reveals distinct and parallel peaks, at times when memory traces are usually formed and/or consolidated. The coordinated rhythmic expression of hippocampal clock gene expression is greatly disordered in mice deficient for the clock gene mPer1, a key player implicated in both, maintenance and adaptative plasticity of circadian clocks. Moreover, Per1‐knockout animals are severely handicapped in a hippocampus‐dependent long‐term spatial learning paradigm. We propose that the dynamics of hippocampal clock gene expression imprint a temporal structure on memory processing and shape at the same time the efficacy of behavioral learning.

Ammonia Induces RNA Oxidation in Cultured Astrocytes and Brain In Vivo

Oxidative stress plays a major role in cerebral ammonia toxicity and the pathogenesis of hepatic encephalopathy (HE). As shown in this study, ammonia induces a rapid RNA oxidation in cultured rat astrocytes, vital mouse brain slices, and rat brain in vivo. Ammonia‐induced RNA oxidation in cultured astrocytes is reversible and sensitive to MK‐801, 1,2‐Bis(o‐aminophenoxy)ethane‐N,N,N′,N′‐tetraacetic acid, apocynin, epigallocatechin gallate, and polyphenon 60, suggesting the involvement of N−methyl−D‐aspartic acid (NMDA) receptor activation, Ca2+, nicotinamide adenine dinucleotide phosphate, and reduced form (NADPH) oxidase‐dependent oxidative stress. Also, hypo‐osmolarity, tumor necrosis factor alpha (TNF‐α), and diazepam increase RNA oxidation in cultured astrocytes, suggesting that the action of different HE‐precipitating factors converges at the level of RNA oxidation. Among the oxidized RNA species, 18S‐rRNA and the messenger RNA (mRNA) coding for the glutamate/aspartate transporter (GLAST) were identified. Cerebral RNA oxidation in acutely ammonia‐loaded rats in vivo is reversible and predominates in neuronal soma and perivascular astrocyte processes. In neuronal dendrites, oxidized RNA colocalizes with the RNA‐binding splicing protein neurooncological ventral antigen (NOVA)‐2 within putative RNA transport granules, which are also found in close vicinity to postsynaptic spines. This indicates that oxidized RNA species may participate in postsynaptic protein synthesis, which is a biochemical substrate for learning and memory consolidation. Neuronal and astroglial RNA oxidation increases also in vital mouse brain slices treated with ammonia and TNF‐α, respectively. Conclusion: Cerebral RNA oxidation is identified as a not yet recognized consequence of acute ammonia intoxication. RNA oxidation may affect gene expression and local protein synthesis and thereby provide another link between reactive oxygen species (ROS)/reactive nitrogen oxide species (RNOS) production and ammonia toxicity. (HEPATOLOGY 2008.)

Orexin (hypocretin)/dynorphin neurons control GABAergic inputs to tuberomammillary neurons

High activity of the histaminergic neurons in the tuberomammillary (TM) nucleus increases wakefulness, and their firing rate is highest during waking and lowest during rapid eye movement sleep. The TM neurons receive a prominent innervation from sleep‐active γ‐aminobutyric acidergic (GABAergic) neurons in the ventrolateral preoptic nucleus, which inhibits them during sleep. They also receive an excitatory input from the orexin‐ and dynorphin‐containing neurons in the lateral hypothalamus, which are critically involved in sleep regulation and whose dysfunction causes narcolepsy. We have used intracellular recordings and immunohistochemistry to study if orexin neurons exert control over the GABAergic inputs to TM neurons in rat hypothalamic slices. Dynorphin suppressed GABAergic inputs and thus disinhibits the TM neurons, acting in concert with orexin to increase the excitability of these neurons. In contrast, both orexin‐A and orexin‐B markedly increased the frequency of GABAergic potentials, while co‐application of orexin and dynorphin produced responses similar to dynorphin alone. Thus, orexins excite TM neurons directly and by disinhibition, gated by dynorphin. These data might explain some of the neuropathology of narcolepsy.

Orexins/hypocretins cause sharp wave- and θ-related synaptic plasticity in the hippocampus via glutamatergic, gabaergic, noradrenergic, and cholinergic signaling

Orexins (OX), also called hypocretins, are bioactive peptides secreted from glucose-sensitive neurons in the lateral hypothalamus linking appetite, arousal and neuroendocrine-autonomic control. Here, OX-A was found to cause a slow-onset long-term potentiation of synaptic transmission (LTPOX) in the hippocampus of young adult mice. LTPOX was induced at Schaffer collateral-CA1 but not mossy fiber-CA3 synapses, and required transient sharp wave-concurrent population field-burst activity generated by the autoassociative CA3 network. Exogenous long theta-frequency stimulation of Schaffer collateral axons erased LTPOX in intact hippocampal slices but not mini slices devoid of the CA3 region. Pharmacological analysis revealed that LTPOX requires co-activation of ionotropic and metabotropic glutamatergic, GABAergic, as well as noradrenergic and cholinergic receptors. Together these data indicate that OX-A induces a state-dependent metaplasticity in the CA1 region associated with sharp-wave and theta rhythm activity as well as glutamatergic, GABAergic, aminergic, and cholinergic transmission. Thus, orexins not only regulate arousal threshold and body weight but also threshold and weight of synaptic connectivity, providing a molecular prerequisite for homeostatic and behavioral state-dependent control of neuronal plasticity and presumably memory functions.

Orexins/hypocretins control bistability of hippocampal long‐term synaptic plasticity through co‐activation of multiple kinases

Aim:  Orexins/hypocretins (OX/Hcrt) are hypothalamic neuropeptides linking sleep–wakefulness, appetite and neuroendocrine control. Their role and mechanisms of action on higher brain functions, such as learning and memory, are not clear.

Water maze performance, exploratory activity, inhibitory avoidance and hippocampal plasticity in aged superior and inferior learners

In 28‐ to 30‐month‐old rats, in vitro short‐term and long‐term potentiation (STP and LTP) were measured in area CA1 of the hippocampus in seven superior and seven inferior learners, that were selected from a pool of 40 rats based on water maze escape performance over a period of 9 days. The aim was to examine whether levels of STP and LTP could account for group differences in learning of water maze escape, spatial preference and wall (thigmotaxis)‐avoidance and in short‐term retention of an inhibitory avoidance task. There was no significant group difference in open‐field exploration, i.e. the number of rearings. In contrast to expectation, the superior and inferior learners did not differ significantly from each other in levels of STP and LTP. However, variability in escape and spatial learning, but not thigmotaxis‐avoidance learning, was significantly predicted by variability in STP and LTP in the superior group. Also, open‐field exploratory rearings were significantly correlated with STP and LTP as well as with maze escape learning in the superior group. The results show that, in the aged superior group, levels of CA1 STP and LTP coincided with residual water maze escape and spatial preference learning as well as open‐field exploration, i.e. behavioural expressions known to be related to hippocampal functioning, but not with learning to avoid thigmotaxis in the maze. The lack of such correlations in the inferior group may be due to the severe impairment in escape and spatial preference learning and/or the influence of yet unknown third variables on these relationships.