Spinal anaesthesia in neonates and infants: what about the cerebral oxygen saturation?

BACKGROUND: Spinal Anaesthesia (SA) has been firmly established as an efficient and safe technique, with minimal cardio-respiratory disturbance when administered in the neonatal period. Our objective was to assess the haemodynamic consequences of SA in infants, particularly its impact on cerebral perfusion using near-infrared spectroscopy (NIRS)-based cerebral oximetry (rSco2). METHODS: All infants up to 60 weeks' postmenstrual age, whether formerly preterm or not, and undergoing spinal anaesthesia, were enrolled. Haemodynamic data records, rSco2 and mean arterial blood pressure (MAP), were prospectively collected before SA (T0) and every five min for 30 min (T30) after the puncture. Compared with baseline measures, any changes of > 10% in rSco2 and of > 20% in MAP were considered clinically significant. Relative variations of data between T0 and T30 were analysed. RESULTS: Data of 103 infants were analysed. The mean relative changes in rSco2 were -2.25% (97.5% CI [-3.97; -0.5]) at T15, and 0.11% (97.5% CI [-1.67; 1.90]) at T30. No significant variation of rSco2 was recorded. The mean changes in MAP were respectively -13.94% (97.5% CI [-17.74; -10.14]) at T15 and -20.27% (97.5% CI [-24,25; -16.29]) at T30. MAP decrease was statistically and clinically significant 30 min after SA. No correlation between changes in MAP and rSco2 was found. The subgroup analysis did not reveal any effect of added intrathecal clonidine or preterm birth history on these results. CONCLUSIONS: In neonate and infants, SA did not cause clinically significant variation in cerebral oxygen saturation. Despite a significant decrease in MAP, cerebral auto-regulation seems to remain effective in neonates and not altered by spinal anaesthesia.

Expression of nucleoside transporter in freshly isolated neurons and astrocytes from mouse brain.

Nucleoside transporters comprise equilibrative ENT1-4 and concentrative CNT1-3. CNTs transport against an intracellular/extracellular gradient and are essential for transmitter removal, independently of metabolic need. ENT1-4 mediate transport until intracellular/extracellular equilibrium of the transported compound, but are very efficient, when the accumulated nucleoside or nucleobase is rapidly eliminated by metabolism. Most nucleoside transporters are membrane-bound, but ENT3 is mainly intracellular. This study uses freshly isolated neurons and astrocytes from two adult mouse strains. In one transgenic strain the neuronal marker Thy1 was associated with a compound fluorescing at one wavelength, and in the other the astrocytic marker GFAP was associated with a compound fluorescent at a different wavelength. Highly purified astrocytic and neuronal populations (as determined by presence/absence of cell-specific genes) were obtained from these mice by fluorescence-activated cell sorting. In each population mRNA analysis was performed by reverse-transcription polymerase chain reaction. CNT1 was absent in both cell types; all other nucleoside transporters were expressed to at least a similar degree (in relation to applied amount of RNA and to a house-keeping gene) in astrocytes as in neurons. Astrocytic ENT3 enrichment was dramatic, but it was not up-regulated after fluoxetine-mediated increase in DNA synthesis. A comparison with results obtained in cultured astrocytes shows that the latter are generally compatible with the present findings and suggests that many observations obtained in intact tissue, mainly by in situ hybridization (which also determines mRNA expression) may underestimate astrocytic nucleoside transporter expression.

Signalling pathways for transactivation by dexmedetomidine of epidermal growth factor receptors in astrocytes and its paracrine effect on neurons.

BACKGROUND AND PURPOSE: Stimulation of astrocytes by the alpha(2)-adrenoceptor agonist dexmedetomidine, a neuroprotective drug, transactivates epidermal growth factor (EGF) receptors. The present study investigates signal pathways leading to release of an EGF receptor ligand and those activated during EGF receptor stimulation, and the response of neurons to dexmedetomidine and to astrocyte-conditioned medium. EXPERIMENTAL APPROACH: Phosphorylation of ERK(1/2) was determined by western blotting and immunocytochemistry, and phosphorylation of EGF receptors by immunoprecipitation and western blotting. mRNA expression of fos family was measured by RT-PCR. KEY RESULTS: Pertussis toxin (0.2 microg ml(-1)) an inhibitor of betagamma subunit dissociation from Galpha(i) protein, and GF 109203X (500 nM), a protein kinase C inhibitor, abolished ERK(1/2) phosphorylation. PP1 (10 microM), inhibiting Src kinase and GM 6001 (10 microM), an inhibitor of Zn-dependent metalloproteinase, abolished ERK(1/2) phosphorylation by dexmedetomidine (50 nM), but not that by EGF (10 ng ml(-1)), showing Src kinase and metalloproteinase activation during the first stage only; AG 1478 (1 microM), an inhibitor of the EGF receptor tyrosine kinase, abolished ERK(1/2) phosphorylation. Dexmedetomidine-induced EGF receptor phosphorylation was prevented by AG 1478, GM 6001, PP1 and GF 109203X and its induction of cfos and fosB by AG 1478 and by U0126 (10 microM), an inhibitor of ERK phosphorylation, indicating downstream effects of ERK(1/2) phosphorylation. EGF and conditioned medium from dexmedetomidine-treated astrocytes, but not dexmedetomidine itself, induced ERK phosphorylation in primary cultures of cerebellar neurons. CONCLUSIONS AND IMPLICATIONS: Dexmedetomidine-induced transactivation pathways were delineated. Its paracrine effect on neurons may account for its neuroprotective effects.

Depolarization-induced, glutamate receptor-mediated, and transactivation-dependent extracellular-signal regulated kinase phosphorylation in cultured cerebellar granule neurons.

Depolarization of 7-8-day-old mouse cerebellar granule neurons in primary cultures, a glutamatergic preparation, by elevation of the extracellular potassium ion concentration ([K+]e) to 45 mM induces an increase of phosphorylation of extracellular-signal regulated kinase 1 and 2 (ERK1/2) at two time periods: 20 min and 60 min after the [K+]e increase. This effect can be mimicked by 5 min of exposure to 50 microM glutamate, suggesting that ERK1/2 phosphorylation in response to the depolarization is brought about by the resulting glutamate release. This concept is supported by the observation that the K+ -mediated stimulation of phosphorylation at both times is inhibited by MK-801, an NMDA antagonist, and by CNQX, an AMPA/kainate antagonist. These antagonists also inhibit the response to glutamate. Both increases in ERK1/2 phosphorylation are also inhibited by GM 6001 (a metalloproteinase inhibitor, preventing 'shedding' of growth factors), by AG 1478 (a receptor tyrosine kinase inhibitor, preventing epidermal growth factor [EGF] receptor activation), and also partly by heparin (inactivating heparin-binding epidermal growth factor [HB-EGF]), suggesting transactivation of epidermal growth factor receptors (EGFR). Transactivation is an intracellular/extracellular signal transduction pathway in which release from receptor- or depolarization-stimulated cells of EGFR ligand(s) (including HB-EGF), catalyzed by a metalloproteinase, stimulates receptor tyrosine kinases on the same (an autocrine effect) or adjacent (a paracrine effect) cells. The expression of HB-EGF as well as of transforming growth factor-alpha (TGF-alpha), two of the EGFR ligands, in the cells was confirmed by reverse transcription polymerase chain reaction, and the only partial inhibition by heparin suggests that both of these EGFR agonists are involved. Such a transactivation may play a major role in glutamate-mediated signaling and plasticity.

Astrocytic energy metabolism consolidates memory in young chicks.

In a single trial discrimination avoidance learning task, chicks learn to distinguish between beads of two colors, which are dipped in either a strong or weak tasting aversant (methyl anthranilate) to induce strongly-reinforced and weakly-reinforced learning, respectively. Consolidation of strongly-reinforced learning can be prevented by inhibitors of glycolysis, such as 2-deoxyglucose and iodoacetate and by inhibitors of oxidative metabolism and the consolidation of weakly-reinforced learning can be promoted by administration of glucose. In the present study we show that bilateral, intracerebral injection of 30 nmol acetate can act like glucose to consolidate labile memory and to restore memory impaired by 2-deoxyglucose administration. Acetate is a metabolic substrate that feeds into the tricarboxylic acid cycle, it is oxidized in astrocytes, but not in neurones. Our data suggest that effects of glucose administered 15-25 min post-training on memory consolidation are mediated via astrocytes not neurons.

Cell culture studies of the role of elevated extracellular glutamate and K+ in neuronal cell death during and after anoxia/ischemia.

During vascular insults in the brain (stroke) the extracellular concentrations of glutamate and K+ increase. It is well acknowledged that the increase in glutamate contributes to the death of neuronal cells at an earlier time than they would have succumbed to energy deprivation as such, but the origin of the released glutamate is not known and cannot easily be studied in the brain in vivo. We have therefore resorted to cell culture studies which have shown that the neuronal rate of formation of glutamate from glutamine is substantially increased during anoxia, especially in glutamatergic neurons. This increase is further enhanced in the presence of excess K+. Phenylsuccinate, a compound that decreases formation of glutamate from glutamine in glutamatergic neurons, counteracts the increase in glutamate formation and, by doing so, improves cell survival. Astrocytes (glial cells) in neuronal-astrocytic co-cultures to some extent protect against anoxic neuronal damage by accumulating glutamate and thus keeping the extracellular glutamate concentration lower than in isolated neuronal cultures.

Complex roles of glutamate in the Gibbs-Ng model of one-trial aversive learning in the new-born chick.

Glutamate is the most widespread excitatory transmitter in the CNS and is probably involved in LTP, a neural phenomenon which may be associated with learning and memory formation. Intracerebral injection of large amounts of glutamate between 5 min and 2.5 min after passive avoidance learning in young chicks inhibits short-term memory, which occurs between 0 and 10 min post-learning in a three-stage model of memory formation first established by Gibbs and Ng(25) [Physiol. Behav. 23:369-375; 1979]. This effect may be attributed to non-specific excitation. Blockade of glutamate uptake by L-aspartic and beta-hydroxamate also abolishes this stage of memory, provided the drug is administered within 2.5 min of learning. Interference with either production of percursors for transmitter glutamate in astrocytes or with glutamate receptors is also detrimental to memory formation, but the effects appear much later. After its release from glutamatergic neurons, glutamate is, to a large extent, accumulated into astrocytes where it is converted to glutamine, which can be returned to glutamatergic neurons and reutilized for synthesis of transmitter glutamate, and partly oxidized as a metabolic substrate. The latter process leads to a net loss of transmitter glutamate which can be compensated for by de novo synthesis of a glutamate precursor alpha-ketoglutarate (alpha KG) in astrocytes, a process which is inhibited by the astrocyte-specific toxin fluoroacetate (R. A. Swanson, personal communication). Intracerebral injection of this toxin abolishes memory during an intermediate stage of memory processing occurring between 20 and 30 min post-training (50) [Cog. Brain Res, 2:93-102; 1994]. Injection of methionine sulfoximine (MSO), a specific inhibitor of glutamine synthetase, which interferes with the re-supply of transmitter glutamate to neurons by inhibition of glutamine synthesis in astrocytes, has a similar effect. This effect of MSO is prevented by intracerebral injection of glutamate, glutamine, or a combination and alpha KG and alanine. MSO must be administered before learning, but does not interfere with acquisition since short-term memory remains intact. Administration of either the NMDA antagonist AP5, the AMPA antagonist DNQX, or the metabotropic receptor antagonist MCPF, also induces amnesia. Memory loss in each case does not occur until after 70 min post-training, during a protein synthesis-dependent long-term memory stage which begins at 60 min following learning. However, to be effective, AP5 must be administered within 60 s following learning, MCPG before 15 min post-learning, and DNQX between 15 and 25 min after learning. Together, these findings suggest that learning results in an immediate release of glutamate, followed by a secondary release of this transmitter at later stages of processing of the memory trace, and that one or both of these increases in extracellular glutamate concentration are essential for the consolidation of long-term memory. Since both fluoroacetate and MSO act exclusively on glial cells, the findings also show that neuronal-glial interactions are necessary during the establishment of memory.

Astrocyte-neuron interaction during one-trial aversive learning in the neonate chick.

During two specific stages of the Gibbs-Ng model of one-trial aversive learning in the neonate chick, we have recently found unequivocal evidence for a crucial involvement of astrocytes. This evidence is metabolic (utilization of the astrocyte-specific energy store, glycogen, during normal learning and inhibition of memory formation by the astrocyte specific metabolic inhibitors, fluoroacetate and methionine sulfoximine) as well as physiological (abolition of memory formation in the presence of ethacrynic acid, an astrocyte-specific inhibitor of cellular reaccumulation of potassium ions). These findings are discussed in the present review in the framework of a more comprehensive description of metabolic and physiological neuronal-astrocytic interactions across an interstitial (extracellular) space bounded by minute processes from either cell type.

Intense furosemide-sensitive potassium accumulation in astrocytes in the presence of pathologically high extracellular potassium levels.

An intense K+ accumulation in primary cultures of astrocytes, occurring when external K+ was increased from 5.4 to 54 mM, was investigated. This increase resulted in a doubling of the K+ content within 10 s. Thirty percent of the accumulation was inhibited by furosemide (2 mM). This drug had no effect on the unidirectional influx of K+ at 5.4 mM K+, but when the extracellular K+ concentration was increased, there appeared to be a furosemide-sensitive component of the influx. This component increased with increasing external K+ levels, reaching 44% of the total influx at 72 mM. These results show that astrocytes exhibit an intense furosemide-sensitive K+ accumulation which is activated by K+ levels resembling those occurring in the extracellular compartment during pathological events. Previous studies on a furosemide-sensitive Cl- pump in cultured astrocytes suggest that this accumulation might be via KCl cotransport, which in other systems is involved in volume control.

Mitochondrial heterogeneity in the brain at the cellular level.

Classically, compartmentation of glutamate metabolism in the brain is associated with the fact that neurons and glia exhibit distinct differences with regard to metabolism of this amino acid. The recent use of 13C-labeled compounds to study this metabolism in conjunction with the availability of cell type-specific tissue culture modes has led to the notion that such compartmentation may even be present in individual cell types, neurons as well as glia. To better understand and explain this, it is proposed that mitochondrial heterogeneity may exist resulting in tricarboxylic acid cycles with different properties regarding cycling rates and ratio as well as coupling to amino acid biosynthesis, primarily involving glutamate and aspartate. These hypotheses are evaluated in the light of current knowledge about mitochondrial structure and function.

Dexmedetomidine-induced stimulation of glutamine oxidation in astrocytes: a possible mechanism for its neuroprotective activity.

Dexmedetomidine is a highly specific alpha2-adrenergic agonist, which is used clinically as an anesthetic adjuvant and in animal experiments has a neuroprotective effect during ischemia. The current study showed that dexmedetomidine enhances glutamine disposal by oxidative metabolism in astrocytes. This effect occurs at pharmacologically relevant concentrations. It is exerted on alpha2-adrenergic receptors and not on imidazoline-preferring sites, and it is large enough to reduce the availability of glutamine as a precursor of neurotoxic glutamate.

Does endogenous norepinephrine regulate potassium homeostasis and metabolism in rat cerebral cortex?

The role of endogenous cerebral norepinephrine (NE) as a modulator of transmembrane cation transport and energy metabolism was evaluated by monitoring extracellular potassium ion activity ([K+[o) in vivo and by measuring cortical Na+,K+-ATPase activity and oxygen consumption in vitro. Ipsilateral cortical NE was depleted by unilateral 6-hydroxydopamine (6-OHDA) lesions of the locus ceruleus (LC). The contralateral cortex was used for control measurements. NE depletion had no effect on resting levels of cortical [K+[o or on the rate of K+ removal from the extracellular space following direct cortical stimulation. There was also no effect of NE depletion on Na+,K+-ATPase activity in cortical homogenates nor on oxygen consumption of cortical slices over a wide range of K+ concentrations. These results indicate tht central NE depletion does not influence movements of cortical K+ either directly through an influence on Na+,K+-ATPase activity or indirectly through effects on oxidative metabolism. It is probable, therefore, that previously described effects of NE on cortical oxidative metabolism are mediated through changes in cerebral perfusion and/or modification of substrate availability in vivo.

Astrocytic adrenoceptors: a major drug target in neurological and psychiatric disorders?

Considerable attention has recently been paid to astrocyte functions, which are briefly summarized. A large amount of data is available about adrenoceptor expression and function in astrocytes, some of it dating back to the 1970's and some of it very recent. This material is reviewed in the present paper. The brain is innervated by noradrenergic fibers extending from locus coeruleus in the brain stem, which in turn is connected to a network of adrenergic and noradrenergic nuclei in the medulla and pons, contributing to the control of (nor)adrenergic, serotonergic, dopaminergic and cholinergic function, both in the central nervous system (CNS) and in the periphery. In the CNS astrocytes constitute a major target for noradrenergic innervation, which regulates morphological plasticity, energy metabolism, membrane transport, gap junction permeability and immunological responses in these cells. Noradrenergic effects on astrocytes are essential during consolidation of episodic, long-term memory, which is reinforced by beta-adrenergic activation. Glycogenolysis and synthesis of glutamate and glutamine from glucose, both of which are metabolic processes restricted to astrocytes, occur at several time-specific stages during the consolidation. Astrocytic abnormalities are almost certainly important in the pathogenesis of multiple sclerosis and in all probability contribute essentially to inflammation and malfunction in Alzheimer's disease and to mood disturbances in affective disorders. Noradrenergic function in astrocytes is severely disturbed by chronic exposure to cocaine, which also changes astrocyte morphology. Development of drugs modifying noradrenergic receptor activity and/or down-stream signaling is advocated for treatment of several neurological/psychiatric disorders and for neuroprotection. Astrocytic preparations are suggested for study of mechanism(s) of action of antidepressant drugs and pathophysiology of mood disorders.

Effects of barbiturates on energy metabolism by cultured astrocytes and neurons in the presence of normal and elevated concentrations of potassium.

Rates of consumption of oxygen and of the formation of CO2 from [U-14C]glucose were studied in primary cultures of either astrocytes or neurons of the cerebral cortex. An increase of the extracellular concentration of potassium from 5 to 55 mM caused an increase in uptake of oxygen and in the production of CO2 in the astrocytes but not in the neurons. Pentobarbital (0.25-1.0 mM) or phenobarbital (1 mM) abolished the stimulation of oxygen uptake and/or the production of CO2 produced by potassium in the astrocytes with less (CO2 production) or no (oxygen uptake) effect at a normal concentration of potassium. In the neurons, pentobarbital (0.1-1.0 mM) caused, in contrast, a moderate inhibition of the production of CO2 and uptake of oxygen which was at least as pronounced at the small concentration of potassium. These results suggest that the pronounced inhibition of the stimulation of uptake of oxygen induced by potassium in brain slices is exerted on astrocytes, whereas the more modest decrease in uptake of oxygen at a small concentration of potassium is a neuronal phenomenon.

Dexmedetomidine, a potent and highly specific alpha 2 agonist, evokes cytosolic calcium surge in astrocytes but not in neurons.

Dexmedetomidine is an extremely potent alpha 2 adrenoceptor agonist which can reduce anaesthetic requirements by up to 90%. However, the precise cellular mechanism of action of this agent is not known. Primary cultures of murine neurons and astrocytes were grown to test the hypothesis that changes in free intracellular calcium may trigger cellular responses to this drug. In astrocyte cultures, experiments revealed a pronounced increase in cytosolic calcium concentration when 100 nM dexmedetomidine was administered. However, in cultured cerebellar granule cell neurons there was no calcium response observed with similar or even higher concentrations of the drug. Results suggest dexmedetomidine may exert its primary effect on the central nervous system via astrocytes.

Barium-induced inhibition of K+ transport mechanisms in cortical astrocytes--its possible contribution to the large Ba2+-evoked extracellular K+ signal in brain.

Homogenous mouse astrocytes in primary cultures were used to investigate the action of different Ba2+ concentrations on 42K transport, membrane potential and Na+,K+-adenosine triphosphatase activity. Five millimolar Ba2+ reduced total K+ influx and efflux (each by 83%) and ouabain-sensitive net K+ uptake (by 80%); it decreased the K+ content, depolarized the membrane potential reversibly and completely inhibited the Na+,K+-adenosine triphosphatase activity. The concentration dependence of these effects was biphasic. Concentrations between 2 and 20 microM affected only the passive K+ fluxes (IC50: 6 microM). Concentrations between 50 microM and 5 mM inhibited the Na+,K+-adenosine triphosphatase and had no further effect on passive fluxes, but inhibited the ouabain-sensitive net uptake of K+ (IC50: 3.1-0.6 mM). It is suggested that the large evoked extracellular K+ increase in the brain observed in Ba2+-treated preparations in vivo or in brain slices to a large extent is due to the impairment of passive and active K+ clearance by glial cells.

What do retinal müller (glial) cells do for their neuronal 'small siblings'?

Müller (radial glial) cells are the predominant glia of the vertebrate retina. They arise, together with rod photoreceptor cells, bipolar cells, and a subset of amacrine cells, from common precursor cells during a late proliferative phase. One Müller cell and a species-specific number of such neurons seem to form a columnar unit within the retinal tissue. In contrast, 'extracolumnar neurons' (ganglion cells, cone photoreceptor cells, horizontal cells, and another subset of amacrine cells) are born and start differentiation before most Müller cells are generated. It may be essential for such neurons to develop metabolic capacities sufficient to support their own survival, whereas late-born ('columnar') neurons seem to depend on a nursing function of their 'sisterly' Müller cell. Thus, out of the cell types within a retinal column it is exclusively the Müller cell that possesses the enzymes for glycogen metabolism. We present evidence that Müller cells express functional insulin receptors. Furthermore, isolated Müller cells rapidly hydrolyse glycogen when they are exposed to an elevated extracellular K+ ion concentration, a signal that is involved in the regulation of neuronal-glial metabolic cooperation in the brain. Müller cells are also thought to be essential for rapid and effective retinal K+ homeostasis. We present patch-clamp measurements on Müller cells of various vertebrate species that all demonstrate inwardly rectifying K+ channels; this type of channel is well-suited to mediate spatial buffering currents. A mathematical model is presented that allows estimation of Müller cell-mediated K+ currents. A simulation analysis shows that these currents greatly limit lateral spread of excitation beyond the borders of light-stimulated retinal columns, and thus help to maintain visual acuity.

Acute and chronic effects of lithium in therapeutically relevant concentrations on potassium uptake into astrocytes.

Potassium uptake into astrocytes in primary cultures was measured by the aid of 42K. Acute application of lithium in concentrations of 1 and 5 mM, but not 0.5 und 0.25 mM, exerted a significant inhibition of the potassium uptake rates. This effect is due to a partial impairment of the ouabain-sensitive part of the uptake into the cells caused by a lithium interaction with the extracellular K+-activated site of the Na+, K+-ATPase. After 14 days of exposure of the astrocytes to 1 mM lithium, the potassium uptake remained lower in the presence of lithium than in its absence. However, the cells had adjusted to the chronic presence of lithium by increasing their potassium uptake to such an extent that, during the exposure to 1 mM lithium, it was indistinguishable from that in cultures from the same batches grown in the absence of lithium and measured in the absence of this compound. The interference by lithium with potassium uptake into astrocytes may well be related to the inhibition of potassium clearance in the CNS described in the literature.

Does the 'mystery of the extra glucose' during CNS activation reflect glutamate synthesis?

The hypothesis is proposed that the repeatedly demonstrated rise in local cerebral blood flow and glucose utilisation during neuronal activation, without a corresponding increase in oxygen utilisation, may reflect glutamine formation from glucose, followed by complete oxidative degradation of glutamate along complex and extended, but well described pathways, known to operate in the brain. The former process requires large amounts of glucose but little oxygen. The latter utilises oxygen but no additional glucose, is a prolonged process and so at any one time involves only a small increase in the rate of oxygen utilisation which is difficult to demonstrate experimentally.

Signaling and gene expression in the neuron-glia unit during brain function and dysfunction: Holger Hydén in memoriam.

Holger Hydén demonstrated almost 40 years ago that learning changes the base composition of nuclear RNA, i.e. induces an alteration in gene expression. An equally revolutionary observation at that time was that a base change occurred in both neurons and glia. From these findings, Holger Hydén concluded that establishment of memory is correlated with protein synthesis, and he demonstrated de novo synthesis of several high-molecular protein species after learning. Moreover, the protein, S-100, which is mainly found in glial cells, was increased during learning, and antibodies towards this protein inhibited memory consolidation. S-100 belongs to a family of Ca(2+)-binding proteins, and Holger Hydén at an early point realized the huge importance of Ca(2+) in brain function. He established that glial cells show more marked and earlier changes in RNA composition in Parkinson's disease than neurons. Holger Hydén also had the vision and courage to suggest that "mental diseases could as well be thought to depend upon a disturbance of processes in glia cells as in the nerve cells", and he showed that antidepressant drugs cause profound changes in glial RNA. The importance of Holger Hydén's findings and visions can only now be fully appreciated. His visionary concepts of the involvement of glia in neurological and mental illness, of learning being associated with changes in gene expression, and of the functional importance of Ca(2+)-binding proteins and Ca(2+) are presently being confirmed and expanded by others. This review briefly summarizes highlights of Holger Hydén's work in these areas, followed by a discussion of recent research, confirming his findings and expanding his visions. This includes strong evidence that glial dysfunction is involved in the development of Parkinson's disease, that drugs effective in mood disorders alter gene expression and exert profound effects on astrocytes, and that neuronal-astrocytic interactions in glutamate signaling, NO synthesis, Ca(2+) signaling, beta-adrenergic activity, second messenger production, protein kinase activities, and transcription factor phosphorylation control the highly programmed events that carry the memory trace through the initial, signal-mediated short-term and intermediate memory stages to protein synthesis-dependent long-term memory.