Ischemia-induced changes in α-tubulin and β-actin mRNA in the gerbil brain and effects of bifemelane hydrochloride

Using in situ hybridization histochemistry, we examined changes in the cytoskeletal protein α-tubulin and β-actin mRNAs in the gerbil brain 14 days after transient ischemia. In an attempt to identify the changes induced in the synthesis of cytoskeletal protein by schemia, we also evaluated the effects of post-ischemia administration of bifemelane on these cytoskeletal proteins. α-Tubulin and α-actin mRNAs were decreased in the CA1 region 14 days after transient ischemia. These decreases coincided with the loss of CA1 pyramidal cells, suggesting that they may have been related to delayed neuronal death. The β-actin mRNA level in ischemic controls was significantly increased in the dentate gyrus, habenular nucleus, and medial and lateral thalamic nuclei, where some afferent nerves project into the hippocampal pyramidal cells. The increased β-actin mRNA suggests that there may be a compensatory enhancement of actin synthesis in the afferent neurons that restores loosened synaptic connections with the ischemic cells in the CA1-4 fields. Administration of bifemelane just after recirculation prevented most of the ischemia-induced mRNA reductions in the CA1 field. Bifemelane's effect may be related to inhibition of Ca²⁺ influx and its radical scavenging activity. When bifemelane was administered to the ischemic group, α-tubulin mRNA levels significantly increased in the dentate gyrus and amygdaloid nucleus, and β-actin mRNAs showed a tendency to increase in the CA3 and CA4 fields, dentate gyrus, and medial and lateral thalamic nuclei. These findings suggest that bifemelane may enhance synthesis of cytoskeletal protein, especially in the ischemic brain, inducing axon outgrowth or synapse formation.     

Expression of mRNA species encoding heat shock protein 90 (hsp90) in control and hyperthermic rabbit brain

Northern blot and in situ hybridization were employed to investigate regional and cell type differences in the expression of hsp90 mRNA species in control and hyperthermic rabbit brain. Riboprobes specific to hsp90 α and β mRNA species were utilized in time-course Northern blot studies on cerebral hemispheres and the cerebellum. Following hyperthermia, levels of hsp90 α and β mRNA were elevated in both brain regions; however, the magnitude of induction was more robust in the cerebellum than in cerebral hemispheres. The pattern of expression of hsp90 genes in rabbit brain was analyzed by in situ hybridization. These studies revealed that hsp90 genes are preferentially expressed in neuronal cell populations in the unstressed mammalian brain. The distribution of hsp90 α and β mRNA was similar, though the signal for the latter was stronger. Following hyperthermia, changes were not detected in the pattern of hsp90 β mRNA expression in the hippocampus. In the cerebellum, a rapid induction of hsp90 β mRNA was apparent in the neuron-enriched granule cell layer, followed by a delayed accumulation in Purkinje neurons. Unlike hsp70, induction of hsp90 was not detected in glial cells of hyperthermic rabbit brain. The localization of hsp90 to neurons suggests that this heat shock protein plays an important role in neuronal function. © 1996 Wiley-Liss, Inc.     

Expression of Ca-dependent (classical) PKC mRNA isoforms after transient cerebral ischemia in gerbil hippocampus

Cerebral ischemia is known to modify the expression of genetic information in the brain. To complement this knowledge, in the present study we have estimated the expression of calcium- and phospholipid-dependent (classical) protein kinase C (c PKC) isoform mRNAs (α, βI and γ) at different time following ischemia. Forebrain cerebral ischemia was performed on Mongolian gerbils by 5 minutes bilateral occlusion of common carotid arteries. At the pointed time the cytoplasmic RNA was extracted from hippocampus and the expression of PKC mRNA quantified by RT PCR technique using GAPDH expression as an internal standard. Results indicate that only one γ isoform of cPKC mRNA expression becomes significantly modified in postischemic hippocampus. A transient increase up to 145% of control within the first 3 h was followed by its decline to 60–65% at a longer recirculation period. This lowered levels returned back to control at 72 h postischemic recovery. This result indicates that γ PKC could be particularly sensitive to ischemic insult and would react in accordance with the other early signals determining ischemic outcome.     

The SCG10-related gene family in the developing rat retina: persistent expression of SCLIP and stathmin in mature ganglion cell layer

Neuronal growth-associated proteins (GAPs), such as GAP-43 and SCG10, are thought to play crucial roles in both axonal and dendritic outgrowth during neural development and regeneration, although the underlying mechanisms remain largely unknown. The recent finding that SCG10 is a microtubule regulator and also the identification of RB3 and SCLIP as two new SCG10-related members prompted us to investigate the roles of SCG10-related family in neural development, using the retina as a model system. We determined the temporal expression and the spatial distribution of SCG10-related mRNAs in the developing rat retina. Semiquantitative analysis by RT-PCR revealed that in prenatal retina, levels of SCG10 and stathmin mRNAs were higher than those of RB3 and SCLIP. In the postnatal retina, the level of SCLIP increased, whereas the level of RB3 remained low. In situ hybridization revealed that GAP-43 and all of the SCG10-related family mRNAs were present in the retinal ganglion cells (RGCs) at all stages of retinal development, and that stathmin mRNA was present in mitotic neuroblastic cells. Differential expression of SCG10 and other members of the family became more evident as retinal development proceeded; SCG10 and RB3 expression were relatively specific in the RGCs and amacrine cells, whereas SCLIP was also evident in bipolar and horizontal cells. Stathmin mRNA was highly expressed both in the RGCs and other interneurons. These results indicate that multiple SCG10-related proteins are expressed in single neurons including RGCs, and suggest that these nGAPs play similar but distinct roles in differentiation and functional maintenance of retinal neurons.     

Restraint stress induces beta-amyloid precursor protein mRNA expression in the rat basolateral amygdala

Several studies have shown the involvement of beta-amyloid precursor proteins (APP) isoforms in physiological process like development of the central nervous system (CNS), functional roles in mature brain, and in pathological process like Alzheimer's disease, neuronal experimental damage, and stress, among others. However, the APP functions are still not clear. In the brain, APP(695) isoform is predominantly found in neurons while APP(751/770) isoforms are predominantly found in astroglial cells and have been associated to neurodegenerative processes. Acute or chronic stress in rats may trigger specific response mechanisms in several brain areas such as amygdala, hippocampus and cortex with the involvement of multiple neurotransmitters. Chronic stress may also induce neuronal injury in rat hippocampus. In situ hybridization (ISH) was used to investigate the expression of APP(695) and APP(751/770) mRNA in amygdala and hippocampus of male Wistar rats (n=4-6 per group) after acute (2 or 6h) or chronic (2h daily/7 days or 6h daily/21 days) restraint stress. Only the APP(695) mRNA expression was significantly increased in the basolateral amygdaloid nuclei following acute or chronic restraint. No APP isoform changed in hippocampus after any stress condition. These results suggest that restraint stress induces changes in gene expression of APP(695) in basolateral amygdaloid nucleus, an area related to stress response.     

Spiny calretinin-immunoreactive neurons in the hilus and CA3 region of the rat hippocampus: Local axon circuits,synaptic connections,and glutamic acid decarboxylase 65/67 mRNA expression

We have used the Golgi method and Golgi electron microscopic techniques to analyze the axonal arborization and efferent connections of spiny calretinin-immunoreactive neurons in the CA3 region and hilus of the rat hippocampal formation. In the hilus, the axons of spiny calretinin-immunoreactive neurons sent out numerous collaterals that arborized in the hilar region and the molecular layer. In the CA3 region, these axons extended mainly to the stratum radiatum and pyramidal layer but also to the stratum oriens and stratum lacunosum-moleculare. Axonal varicosities were distributed widely throughout the axonal collaterals. Electron microscopic studies revealed that the axon terminals of spiny calretinin-immunoreactive neurons established synaptic contacts mainly with dendritic shafts. We next analyzed the expression of glutamic acid decarboxylase (GAD65/67) mRNAs in spiny nonpyramidal neurons that were identified by calretinin immunoreactivity. We found that spiny calretinin-positive neurons in the CA3 region and hilus of the rat hippocampal formation expressed the two isoforms of GAD: GAD65 and GAD67 mRNAs. These findings show that the spiny calretinin-immunoreactive neurons of hippocampus give rise to local axonal arborizations, suggesting that they are inhibitory. J. Comp. Neurol. 404:438–448, 1999. © 1999 Wiley-Liss, Inc.     

GABAA Receptor subunits have differential distributions in the rat retinae: In situ hybridization and immunohistochemistry

The distributions of nine different subunits of the gamma-aminobutyric acid_A (GABA_A) receptor (α₁, α₂, α₃, ₅; β₁, β₂, β₃; γ₂; δ) were investigated in the rat retina using immunocytochemistry and in situ hybridization. With the exception of the α₅ subunit, all subunits could be localized. Each subunit was expressed in characteristic strata within the inner plexiform layer (IPL). Some subunits (e.g., γ₂) showed a ubiquitous distribution, while others (e.g., δ) were restricted to narrow sublayers. Double labeling experiments using different combinations of the subunit-specific antibodies revealed colocalizations of subunits within individual neurons. Additionally, GABA_A receptor subunits were mapped to distinct populations of retinal neurons by coapplication of defined immunocytochemical markers and subunit-specific antibodies. Cholinergic amacrine cells were found to express the α₂, β1, β_2/3 and δ subunits, while dopaminergic amacrine cells express the α₂, α₃ and γ₂ subunits. Dissociated rod bipolar cells express the γ₂ subunits. In summary, this study provides evidence for the existence of multiple GABA_A receptor subtypes in the retina. The distinct stratification pattern of the subunits in the IPL suggests that different functional circuits involve specific subtypes of GABAA receptors. © 1995 Wiley-Liss, Inc.     

Cell-specific expression of type II calcium/calmodulin-dependent protein kinase isoforms and glutamate receptors in normal and visually deprived lateral geniculate nucleus of monkeys

In situ hybridization histochemistry and immunocytochemistry were used to map distributions of cells expressing mRNAs encoding α, β, γ, and δ isoforms of type II calcium/calmodulin-dependent protein kinase (CaMKII), α-amino-3-hydroxy-5-methyl-4-isoxazoleproprionate (AMPA)/kainate receptor subunits, (GluR1–7), and N-methyl-D-aspartate (NMDA) receptor subunits, NR1 and NR2A-D, or stained by subunit-specific immunocytochemistry in the dorsal lateral geniculate nuclei of macaque monkeys. Relationships of specific isoforms with particular glutamate receptor types may be important elements in neural plasticity. CaMKII-α is expressed only by neurons in the S laminae and interlaminar plexuses of the dorsal lateral geniculate nucleus, but may form part of a more widely distributed matrix of similar cells extending from the geniculate into adjacent nuclei. CaMKII-β, -γ, and -δ isoforms are expressed by all neurons in principal and S laminae and interlaminar plexuses. In principal laminae, they are down-regulated by monocular deprivation lasting 8–21 days. All glutamate receptor subunits are expressed by neurons in principal and S laminae and interlaminar plexuses. The AMPA/kainate subunits, GluR1, 2, 5, and 7, are expressed at low levels, although GluR1 immunostaining appears selectively to stain interneurons. GluR3 is expressed at weak, GluR 6 at moderate and GluR 4 at high levels. NMDA subunits, NR1 and NR2A, B, and D, are expressed at moderate to low levels. GluR4, GluR6 and NMDA subunits are down-regulated by visual deprivation. CaMKII-α expression is unique in comparison with other CaMKII isoforms which may, therefore, have more generalized roles in cell function. The results demonstrate that all of the isoforms are associated with NMDA receptors and with AMPA receptors enriched with GluR4 subunits, which implies high calcium permeability and rapid gating. J. Comp. Neurol. 390:278–296, 1998. © 1998 Wiley-Liss, Inc.     

Localization of cytochrome oxidase (COX) activity and COX mRNA in the hippocampus and entorhinal cortex of the monkey brain: correlation with specific neuronal pathways

Cytochrome oxidase (COX) activity and COX II mRNA expression were localized in the hippocampal formation and entorhinal cortex of the rhesus monkey brain by means of enzyme histochemistry and in situ hybridization, respectively. Within the hippocampal formation, the terminal field of the perforant pathway showed the highest levels of COX activity, whereas COX II mRNA was localized mainly in neuronal cell bodies. In the entorhinal cortex. COX II mRNA was detected in neuronal cell bodies of layers II and IV. These results indicate that the pattern of localization of COX and its mRNA in entorhinal cortex correlates with the input and output pathways of the hippocampus.     

α1-Adrenergic receptors in the brain: characterization in astrocytic glial cultures and comparison with neuronal cultures

Binding of [¹²⁵I]HEAT to membranes prepared from primary cultures of astrocytic glial cells was time-dependent and 70–85% specific. Various adrenergic agonists and antagonists competed for [¹²⁵I]HEAT binding to the potencies of prazosin >, yohimbine , clonidine, norepinephrine (NE), and propranolol. Scatchard analysis showedB_max of 209 fmol/mg protein and a K_d of 184 pM for [¹²⁵I]HEAT binding by astrocytic glial membranes. Pretreatment of astrocytes with NE resulted in a dose-dependent downregulation of [¹²⁵I]HEAT binding sites with a maximal response observed after 8 h at 100 μM NE. Removal of NE from cultures after pretreatment resulted in a time- and protein synthesis-dependent recovery of binding sites to control levels within 120 h. Incubation of astrocytic glial cultures with NE stimulated phosphoinositide (PI) hydrolysis in a time- and dose-dependent manner with a maximal stimulation of 2-fold observed in 60 min by 100 μM NE.Clonidine expressed differential effects on α₁-adrenergic receptors of the neuronal and astrocytic glial cultures. Pretreatment with 10 μM clonidine caused a 40% decrease in the B_max of [¹²⁵I]HEAT binding without influencing the K_d value in neuronal cultures. This downregulatory effect of clonidine was associated with a reduction in the ability of NE to stimulate PI hydrolysis in clonidine pretreated cells. In contrast to neuronal cultures, clonidine neither downregulated [¹²⁵I]HEAT binding sites nor stimulated PI hydrolysis in glial cultures.     

Distribution of mRNAs encoding transforming growth factors‐β1, ‐2, and ‐3 in the intact rat brain and after experimentally induced focal ischemia

Transforming growth factors‐β1 (TGF‐β1), ‐2, and ‐3 form a small group of related proteins involved in the regulation of proliferation, differentiation, and survival of various cell types. Recently, TGF‐βs were also demonstrated to be neuroprotective. In the present study, we investigated their distribution in the rat brain as well as their expression following middle cerebral artery occlusion. Probes were produced for all types of TGF‐βs, and in situ hybridization was performed. We demonstrated high TGF‐β1 expression in cerebral cortex, hippocampus, central amygdaloid nucleus, medial preoptic area, hypothalamic paraventricular nucleus, substantia nigra, brainstem reticular formation and motoneurons, and area postrema. In contrast, TGF‐β2 was abundantly expressed in deep cortical layers, dentate gyrus, midline thalamic nuclei, posterior hypothalamic area and mamillary body, superior olive, areas of monoaminergic neurons, spinal trigeminal nucleus, dorsal vagal complex, cerebellum, and choroid plexus, and a high level of TGF‐β3 mRNA was found in cerebral cortex, hippocampus, basal amygdaloid nuclei, lateral septal nucleus, several thalamic nuclei, arcuate and supramamillary nuclei, superior colliculus, superior olive, brainstem reticular formation and motoneurons, area postrema, and inferior olive. Focal brain ischemia induced TGF‐βs with markedly different expression patterns. TGF‐β1 was induced in the penumbral region of cortex and striatum, whereas TGF‐β2 and ‐β3 were induced in different layers of the ipsilateral cortex. The expression of the subtypes of TGF‐βs in different brain regions suggests that they are involved in the regulation of different neurons and bind to different latent TGF‐β binding proteins. Furthermore, they might have subtype‐specific functions following ischemic attack. J. Comp. Neurol. 518:3752–3770, 2010. © 2010 Wiley‐Liss, Inc.     

Electrophysiological actions of norepinephrine in rat lateral hypothalamus. II. An in vitro study of the effects of iontophoretically applied norepinephrine on LH neuronal responses to γ-aminobutyric acid (GABA)

The preceding studies demonstrated that norepinephrine (NE) can consistently augment synaptically mediated (70%) and γ-aminobutyric acid (GABA)-induced (69%) inhibitory responses of lateral hypothalamic (LH) neurons in vivo. The present experiments further characterized the interactions of NE with LH neuronal responses to GABA in terms of α- and ß-receptor mechanisms and demonsrated the utility of the in vitro LH tissue slice preparation as a model for future extra- and intracellular studies of NE modulatory phenomena. Extracellular activity of LH cells was recorded from diencephalic slices (450 μm) incubated in artificial cerebrospinal fluid at 33 °C. Interactions between iontophoretically applied NE, isoproterenol (ISO) or phenylephrine (PE) and responses of LH neurons (n = 64) to GABA microiontophoresis were quantitated and characterized using computer-generated ratemeter and histogram records. This analysis revealed two distinct actions of NE on GABA-induced responses of LH neurons. In 8 of 32 cells tested (25%), locally applied NE markedly enhanced inhibitory responses to GABA iontophoresis in a manner identical to that observed in vivo. However, in 20 cells (62.5%), iontophoretic application of NE produced a clear antagonism of GABA responses. NE also exerted dual effects on the background firing rate of LH neurons, causing both inhibition and excitation. Overall, in those cells where NE administration increased spontaneous discharge, it either antagonized or had no effect on GABA-mediated inhibition. In contrast, spontaneous firing rate was never elevated above control levels in those cases where NE potentiated GABA responses. Additional experiments demonstrated that the GABA potentiating actions of the benzodiazepine, flurazepam, were preserved in LH tissue slice preparations. In addition, iontophoretic application of the ß-agonist, ISO, routinely suppressed the spontaneous activity of LH neurons and mimicked the facilitating action of NE on GABA. Likewise, microiontophoretic application of 8-bromo cyclic adenosine monophosphate (AMP) enhanced GABA-induced inhibition of LH firing rate in each of 11 cells tested. On the other hand, local administration of the alpha agonist, PE, routinely produced NE-like antagonism of GABA inhibition along with increases in spontaneous firing rate. Taken together these findings indicate that the commonly observed in vivo phenomena of NE augmentation of GABA and suppression of LH neuron spontaneous firing can be demonstrated in vitro, and most likely result from activation of beta adrenoceptors and subsequent elevation of cyclic AMP levels. As such these results suggest that the in vitro preparation will be a useful tool for further investigation of the transmembrane and intracellular events associated with noradrenergically mediated enhancement of GABA. However, in contrast to results obtained in vivo, NE antagonism of GABA inhibition and excitatory effects on spontaneous activity were more frequently observed in LH slices and appear to be mediated by alpha receptor activation. The reduced capacity of NE to augment GABA in vitro might be related to changes in the balance between α- and ß-mediated effects rather than deficits in GABA-facilitating mechanisms, since ISO, cyclic AMP and the benzodiazepine were all routinely capable of enhancing GABAergic responses.     

Distribution of β-amyloid and amyloid precursor protein in the brain of spawning (senescent) salmon: a natural, brain-aging model

Brain amyloid precursor protein (APP), a normal constituent of neurons, glial cells and cerebrospinal fluid, has several proposed functions (e.g., in neuronal growth and survival). It appears, however, that altered processing of APP is an initial or downstream step in the neuropathology of brain aging, Alzheimer's disease (AD), and Down's syndrome (DS). Some studies suggest that proteolytic cleavage of APP, producing β-amyloid (Aβ_1–42), could have neurotoxic or neuroprotective effects. In this study, we utilized antibodies to human APP₆₉₅ and Aβ_1–42, and Congo red staining, to search for amyloid deposition in the brain of semelparous spawning kokanee salmon (Oncorhynchus nerka kennerlyi). Intracellular APP₆₉₅ immunoreactivity (APP-ir) was observed in brain regions involved in gustation (glomerulosus complex), olfaction (putative hippocampus, olfactory bulb), vision (optic tectum), the stress response (nucleus preopticus and nucleus lateralis tuberis), reproductive behavior (nucleus preopticus magnocellularis, nucleus preopticus periventricularis, ventral telencephalon), and coordination (cerebellum). Intra- and extra-neuronal Aβ_1–42 immunoreactivity (Aβ-ir) were present in all APP-ir regions except the nucleus lateralis tuberis and Purkinje cells of the cerebellum (coordination). Thus, the relationship between APP and Aβ deposition during brain aging could shed light on the processing of APP into Aβ, neurodegeneration, and possible protection of neurons that are functioning in spawning but senescent salmon. Pacific salmon, with their predictable and synchronized life history, could provide research options not available with the existing models for studies of brain aging and amyloidosis.     

Estrogen and brain-derived neurotrophic factor (BDNF) in hippocampus: complexity of steroid hormone-growth factor interactions in the adult CNS

In the CNS, there are widespread and diverse interactions between growth factors and estrogen. Here we examine the interactions of estrogen and brain-derived neurotrophic factor (BDNF), two molecules that have historically been studied separately, despite the fact that they seem to share common targets, effects, and mechanisms of action. The demonstration of an estrogen-sensitive response element on the BDNF gene provided an impetus to explore a direct relationship between estrogen and BDNF, and predicted that the effects of estrogen, at least in part, might be due to the induction of BDNF. This hypothesis is discussed with respect to the hippocampus, where substantial evidence has accumulated in favor of it, but alternate hypotheses are also raised. It is suggested that some of the interactions between estrogen and BDNF, as well as the controversies and implications associated with their respective actions, may be best appreciated in light of the ability of BDNF to induce neuropeptide Y (NPY) synthesis in hippocampal neurons. Taken together, this tri-molecular cascade, estrogen-BDNF-NPY, may be important in understanding the hormonal regulation of hippocampal function. It may also be relevant to other regions of the CNS where estrogen is known to exert profound effects, such as amygdala and hypothalamus; and may provide greater insight into neurological disorders and psychiatric illness, including Alzheimer's disease, depression and epilepsy.     

Molecular profiling of the developing avian telencephalon: Regional timing and brain subdivision continuities

In our companion study (Jarvis et al. [2013] J Comp Neurol. doi: 10.1002/cne.23404) we used quantitative brain molecular profiling to discover that distinct subdivisions in the avian pallium above and below the ventricle and the associated mesopallium lamina have similar molecular profiles, leading to a hypothesis that they may form as continuous subdivisions around the lateral ventricle. To explore this hypothesis, here we profiled the expression of 16 genes at eight developmental stages. The genes included those that define brain subdivisions in the adult and some that are also involved in brain development. We found that phyletic hierarchical cluster and linear regression network analyses of gene expression profiles implicated single and mixed ancestry of these brain regions at early embryonic stages. Most gene expression–defined pallial subdivisions began as one ventral or dorsal domain that later formed specific folds around the lateral ventricle. Subsequently a clear ventricle boundary formed, partitioning them into dorsal and ventral pallial subdivisions surrounding the mesopallium lamina. These subdivisions each included two parts of the mesopallium, the nidopallium and hyperpallium, and the arcopallium and hippocampus, respectively. Each subdivision expression profile had a different temporal order of appearance, similar in timing to the order of analogous cell types of the mammalian cortex. Furthermore, like the mammalian pallium, expression in the ventral pallial subdivisions became distinct during prehatch development, whereas the dorsal portions did so during posthatch development. These findings support the continuum hypothesis of avian brain subdivision development around the ventricle and influence hypotheses on homologies of the avian pallium with other vertebrates. J. Comp. Neurol. 521:3666–3701, 2013. © 2013 Wiley Periodicals, Inc.     

Altered intracellular localization of the glutamate receptor channel δ2 subunit in weaver and reeler Purkinje cells

The glutamate receptor (GluR) channel δ2 subunit is expressed abundantly and specifically in cerebellar Purkinje cells. Our previous study demonstrated that the GluR δ2 mRNA is expressed as early as embryonic day 15 prior to Purkinje cell synaptogenesis, and its protein product accumulates in dendritic spines during normal Purkinje cell maturation. In this study, we examined expression and distribution of the GluR δ2 in the weaver and reeler mutant cerebelli, which show abnormal cytoarchitecture and neural circuitry. In situ hybridization analysis showed that GluR δ2 mRNA was expressed in entire Purkinje cells in both mutant mice. Immunohistochemical analysis revealed that intracellular localization of GluR δ2 was altered in some region of mutant cerebelli. In the cortical surface where Purkinje cells form synapses with parallel fibers, GluR δ2-immunoreactivity was restricted to dendritic spines of Purkinje cells as observed in normal mice. In contrast, in the subcortical region where granule cells and parallel fibers are absent, the immunoreactivity was found widely in Purkinje dendrites. Thus, the GluR δ2 protein did not accumulate to the dendritic spines of Purkinje cells lacking synaptic contact with parallel fibers. These results suggest that the expression of both GluR δ2 mRNA and protein is independent of abnormalities in the mutant cerebelli, but relocalization of the GluR δ2 protein might depend on the formation of synapses between Purkinje cells and parallel fibers.