Fatal ruxolitinib-related JC virus meningitis.

We report a possible association between ruxolitinib and JC virus meningitis. A 72-year-old man with myelofibrosis started treatment with ruxolitinib. Fourteen days later, the patient presented to the emergency department with fever and nausea. HIV test was negative. Ruxolitinib was suspended. Symptoms progressed with neck stiffness, cognitive impairment, and motor aphasia. CSF was positive for JC virus. MRI showed nonspecific abnormal findings. Five days after the clinical debut, the patient died. The clinical picture, MRI imaging, and positive JC virus PCR in CSF strongly suggest ruxolitinib-related JC virus meningitis.

Selective down-regulation of α4ß2 neuronal nicotinic acetylcholine receptors in the brain of uremic rats with cognitive impairment.

Cognitive impairment is common in patients with chronic kidney disease. Brain nicotinic acetylcholine receptors modulate cognitive functions, such as learning and memory. Pharmacological cholinergic enhancement is useful in patients with cognitive dysfunction. The major nicotinic acetylcholine receptor subtypes in the brain are heteromeric α4ß2 and homomeric α7 receptors. To study the involvement of neuronal acetylcholine receptors in cognitive impairment in uremic rats, bilateral nephrectomy was performed. 24 weeks after nephrectomy, memory was assessed using the one trial step-down inhibitory avoidance test. Neuronal nicotinic acetylcholine receptors in the brain were studied by radioligand binding, immunoprecipitation, Western blot and sucrose gradient experiments. We demonstrated that rats with severe renal failure show disorders of short term memory. Long term memory was not altered in these rats. The number of functional α4ß2 heteromeric neuronal nicotinic receptors was decreased in the brains of rats with severe renal failure. There was a significant correlation between the degree of renal impairment and the number of heteromeric nicotinic acetylcholine receptors in the brain. The down-regulation of functional α4ß2 receptors in the brains of rats with severe renal failure was not due to a reduction of α4 or ß2 subunit proteins. The number of α7 homomeric neuronal nicotinic acetylcholine receptors was not altered. These findings may have important clinical significance for the management of cognitive impairment in patients with chronic kidney disease.

The influence of cold temperature on cellular excitability of hippocampal networks.

The hippocampus plays an important role in short term memory, learning and spatial navigation. A characteristic feature of the hippocampal region is its expression of different electrical population rhythms and activities during different brain states. Physiological fluctuations in brain temperature affect the activity patterns in hippocampus, but the underlying cellular mechanisms are poorly understood. In this work, we investigated the thermal modulation of hippocampal activity at the cellular network level. Primary cell cultures of mouse E17 hippocampus displayed robust network activation upon light cooling of the extracellular solution from baseline physiological temperatures. The activity generated was dependent on action potential firing and excitatory glutamatergic synaptic transmission. Involvement of thermosensitive channels from the transient receptor potential (TRP) family in network activation by temperature changes was ruled out, whereas pharmacological and immunochemical experiments strongly pointed towards the involvement of temperature-sensitive two-pore-domain potassium channels (K(2P)), TREK/TRAAK family. In hippocampal slices we could show an increase in evoked and spontaneous synaptic activity produced by mild cooling in the physiological range that was prevented by chloroform, a K(2P) channel opener. We propose that cold-induced closure of background TREK/TRAAK family channels increases the excitability of some hippocampal neurons, acting as a temperature-sensitive gate of network activation. Our findings in the hippocampus open the possibility that small temperature variations in the brain in vivo, associated with metabolism or blood flow oscillations, act as a switch mechanism of neuronal activity and determination of firing patterns through regulation of thermosensitive background potassium channel activity.

An extracellular RRR motif flanking the M1 transmembrane domain governs the biogenesis of homomeric neuronal nicotinic acetylcholine receptors.

We have previously demonstrated that the highly conserved R209, that flanks the M1 transmembrane segment of nicotinic acetylcholine (ACh) receptors, is required for the transport of assembled homomeric neuronal α7 nicotinic ACh receptors to the cell surface. In the present paper we show that basic residues at positions 208 and 210 are necessary for the assembly of α7 receptors. On the contrary, a basic residue at position 210 of α3 subunit decreases the assembly of heteromeric neuronal α3ß4 nicotinic ACh receptors. A basic residue at position 210 of the ß4 subunit slightly decreases α3ß4 receptor expression. We conclude that a pre-M1 RRR motif is necessary for the biogenesis of homomeric α-bungarotoxin-sensitive neuronal α7 nicotinic ACh receptors.

Postnatal alterations of the inhibitory synaptic responses recorded from cortical pyramidal neurons in the Lis1/sLis1 mutant mouse.

Mutations in the mouse Lis1 gene produce severe alterations in the developing cortex. We have examined some electrophysiological responses of cortical pyramidal neurons during the early postnatal development of Lis/sLis1 mutant mice. In P7 and P30 Lis1/sLis1 neurons we detected a lower frequency and slower decay phase of mIPSCs, and at P30 the mIPSCs amplitude and the action potential duration were reduced. Zolpidem (an agonist of GABAA receptors containing the alpha1 subunit) neither modified the amplitude nor the decay time of mIPSCs at P7 in Lis1/sLis1 neurons, whereas it increased the decay time at P30. The levels of GABAA receptor alpha1 subunit mRNA were reduced in the Lis1/sLis1 brain at P7 and P30, whereas reduced levels of the corresponding protein were only found at P7. These results demonstrate the presence of functional alterations in the postnatal Lis1/sLis1 cortex and point to abnormalities in GABAA receptor subunit switching processes during postnatal development.

Immunohistochemical localization of the voltage-gated potassium channel subunit Kv1.4 in the central nervous system of the adult rat.

A large set of voltage-gated potassium channels is involved in regulating essential aspects of neuronal function in the central nervous system, thus contributing to the ability of neurons to respond to a given input. In the present study, we used immunocytochemical methods to elucidate the regional, cellular and subcellular distribution of the voltage-gated potassium channel subunit Kv1.4, a member of the Shaker subfamily, in the brain. At the light microscopic level, the Kv1.4 subunit showed a unique distribution pattern, being localized in specific neuronal populations of the rat brain. The neuronal regions expressing the highest levels of Kv1.4 protein included the cerebral cortex, the hippocampus, the posterolateral and posteromedial ventral thalamic nuclei, the dorsolateral and medial geniculate nuclei, the substantia nigra and the dorsal cochlear nucleus. The Kv1.4 subunit was also present in other neuronal populations, with different levels of Kv1.4 immunoreactivity. In all immunolabeled regions, the Kv1.4 subunit was mostly diffusely distributed and, to a lesser extent, it stained cell bodies and proximal dendrites. Furthermore, Kv1.4 immunoreactivity was also detected in nerve terminals and axonal terminal fields. At the electron microscopic level, Kv1.4 was located postsynaptically in dendritic spines and shafts at extrasynaptic sites, as well as presynaptically in axon and active zone of axon terminals, in the neocortex and hippocampus. The findings indicate that Kv1.4 channels are widely distributed in the rat brain and suggest that activation of this channel would have different modulatory effects on neuronal excitability.

Role of the large cytoplasmic loop of the alpha 7 neuronal nicotinic acetylcholine receptor subunit in receptor expression and function.

The role of the large intracellular loop of the nicotinic acetylcholine receptor (nAChR) alpha7 subunit in the expression of functional channels was studied. For this purpose, systematic deletions and substitutions were made throughout the loop and the ability of the mutated alpha7 subunits to support expression of functional nAChRs at the Xenopus oocyte membrane was tested. Surface nAChR expression was abolished upon removal of sequences at two regions, a 29-amino acid segment close to the N-terminus of the loop (amino acids 297-325) and adjacent to the third transmembrane region and an 11-amino acid segment near the fourth transmembrane region. Some residues (amino acids 317-322) within the 29 amino acids N-terminal segment could be substituted by others but not deleted without loss of expression, suggesting that a certain structure, determined by the number of amino acids rather than by their identity, has to be maintained in this region. The contiguous sequence M323 K324 R325 did not tolerate deletions and substitutions. Removal of the rest of the cytoplasmic loop was not deleterious; even higher expression levels (2-4-fold) were obtained upon large deletions of the loop (Delta399-432 and Delta339-370). High expression levels were observed provided that a minimal sequence of three amino acids (E371, G372, and M373) was present. In addition, some electrophysiological properties of mutant nAChRs were modified. Substitution of the EGM sequence by other protein segments produced a variety of effects, but, in general, insertions were not well tolerated, suggesting the existence of tight structural restrictions in the large cytoplasmic region of the rat alpha7 subunit.

Transcription factors NF-Y and Sp1 are important determinants of the promoter activity of the bovine and human neuronal nicotinic receptor beta 4 subunit genes.

The beta4 subunit is a component of the neuronal nicotinic acetylcholine receptors which control catecholamine secretion in bovine adrenomedullary chromaffin cells. The promoter of the gene coding for this subunit was characterized. A proximal region (from minus sign99 to minus sign64) was responsible for the transcriptional activity observed in chromaffin, C2C12, and COS cells. Within this region two cis-acting elements that bind transcription factors Sp1 and NF-Y were identified. Mutagenesis of the two elements indicated that they cooperate for the basal transcription activity of the promoter. The human beta4 promoter, that was also characterized, shared structural and functional homologies with the bovine promoter. Thus, two adjacent binding elements for Sp1 and NF-Y were detected. Whereas the Sp1 site was an important determinant of the promoter activity, the NF-Y site may have cell-specific effects. Given that these promoters showed no structural or functional homology with the previously characterized rat beta4 subunit promoter (Bigger, C. B., Casanova, E. A., and Gardner, P. D. (1996) J. Biol. Chem. 271, 32842--32848) except for the involvement of an Sp1 binding element, we propose that constitutive expression of the beta4 subunit gene in these three close species may be controlled by the general transcription factor Sp1. Nevertheless, other components could determine species-specific beta4 subunit expression.