Cortical State Fluctuations across Layers of V1 during Visual Spatial Perception

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Regulation of Ca2+ Release by InsP3 in Single Guinea Pig Hepatocytes and Rat Purkinje Neurons

The repetitive spiking of free cytosolic [Ca²⁺] ([Ca²⁺]_i) during hormonal activation of hepatocytes depends on the activation and subsequent inactivation of InsP₃-evoked Ca²⁺ release. The kinetics of both processes were studied with flash photolytic release of InsP₃ and time resolved measurements of [Ca²⁺]_i in single cells. InsP₃ evoked Ca²⁺ flux into the cytosol was measured as d[Ca²⁺]_i/dt, and the kinetics of Ca²⁺ release compared between hepatocytes and cerebellar Purkinje neurons. In hepatocytes release occurs at InsP₃ concentrations greater than 0.1–0.2 μM. A comparison with photolytic release of metabolically stable 5-thio-InsP₃ suggests that metabolism of InsP₃ is important in determining the minimal concentration needed to produce Ca²⁺ release. A distinct latency or delay of several hundred milliseconds after release of low InsP₃ concentrations decreased to a minimum of 20–30 ms at high concentrations and is reduced to zero by prior increase of [Ca²⁺]_i, suggesting a cooperative action of Ca²⁺ in InsP₃ receptor activation. InsP₃-evoked flux and peak [Ca²⁺]_i increased with InsP₃ concentration up to 5–10 μM, with large variation from cell to cell at each InsP₃ concentration. The duration of InsP₃-evoked flux, measured as 10–90% risetime, showed a good reciprocal correlation with d[Ca²⁺]_i/dt and much less cell to cell variation than the dependence of flux on InsP₃ concentration, suggesting that the rate of termination of the Ca²⁺ flux depends on the free Ca²⁺ flux itself. Comparing this data between hepatocytes and Purkinje neurons shows a similar reciprocal correlation for both, in hepatocytes in the range of low Ca²⁺ flux, up to 50 μM · s⁻¹ and in Purkinje neurons at high flux up to 1,400 μM · s⁻¹. Experiments in which [Ca²⁺]_i was controlled at resting or elevated levels support a mechanism in which InsP₃-evoked Ca²⁺ flux is inhibited by Ca²⁺ inactivation of closed receptor/channels due to Ca²⁺ accumulation local to the release sites. Hepatocytes have a much smaller, more prolonged InsP₃-evoked Ca²⁺ flux than Purkinje neurons. Evidence suggests that these differences in kinetics can be explained by the much lower InsP₃ receptor density in hepatocytes than Purkinje neurons, rather than differences in receptor isoform, and, more generally, that high InsP₃ receptor density promotes fast rising, rapidly inactivating InsP₃-evoked [Ca²⁺]_i transients.     

Basal forebrain cholinergic system in the dementias: Vulnerability,resilience, and resistance

The basal forebrain cholinergic neurons (BFCN) provide the primary source of cholinergic innervation of the human cerebral cortex. They are involved in the cognitive processes of learning, memory, and attention. These neurons are differentially vulnerable in various neuropathologic entities that cause dementia. This review summarizes the relevance to BFCN of neuropathologic markers associated with dementias, including the plaques and tangles of Alzheimer's disease (AD), the Lewy bodies of diffuse Lewy body disease, the tauopathy of frontotemporal lobar degeneration (FTLD-TAU) and the TDP-43 proteinopathy of FTLD-TDP. Each of these proteinopathies has a different relationship to BFCN and their corticofugal axons. Available evidence points to early and substantial degeneration of the BFCN in AD and diffuse Lewy body disease. In AD, the major neurodegenerative correlate is accumulation of phosphotau in neurofibrillary tangles. However, these neurons are less vulnerable to the tauopathy of FTLD. An intriguing finding is that the intracellular tau of AD causes destruction of the BFCN, whereas that of FTLD does not. This observation has profound implications for exploring the impact of different species of tauopathy on neuronal survival. The proteinopathy of FTLD-TDP shows virtually no abnormal inclusions within the BFCN. Thus, the BFCN are highly vulnerable to the neurodegenerative effects of tauopathy in AD, resilient to the neurodegenerative effect of tauopathy in FTLD and apparently resistant to the emergence of proteinopathy in FTLD-TDP and perhaps also in Pick's disease. Investigations are beginning to shed light on the potential mechanisms of this differential vulnerability and their implications for therapeutic intervention.

     

Characteristics of store-operated calcium channels activated by depletion of the endoplasmic reticulum ryanodine-sensitive calcium stores in rat dorsal root ganglion neurons

In neurons of the rat dorsal root ganglia (DRG), using a patch-clamp technique in the whole-cell configuration, we studied the characteristics of calcium channels activated by depletion of the ryanodine-sensitive calcium stores of the endoplasmic reticulum. Current-voltage (I-V) relationships of these store-operated calcium channels were obtained by subtraction of the integral I-V characteristics after application of caffeine from the integral I-V characteristics of calcium channels in the control. Currents through store-operated calcium channels could be induced by application of a series of hyperpolarization current pulses to the cell under conditions of replacement of a calcium-free solution containing caffeine by a caffeine-free solution containing 2 mM Ca²⁺. In this case, the following two main conditions were abserved: Voltage-operated calcium channels were inactivated, while a gradient of the electrochemical potential for calcium ions was increased, which made easier passing of these currents through store-operated calcium channels. Therefore, we found that in DRG neurons, despite the presence of great numbers of both voltage-operated and receptor-dependent calcium channels, one more mechanism underlying the entry of calcium through store-operated channels does exist. Neirofiziologiya/Neurophysiology, Vol. 39, No. 3, pp. 195–200, May–June, 2007.     

Dendrin在初级感觉神经元内的表达

Dendrin作为一类新的树突蛋白,在脑内的表达随着行为活动的改变而变化。最近本课题组采用免疫组织化学方法对dendrin在小鼠初级感觉神经元内的表达进行了研究,并进一步研究外周神经损伤后dendrin在背根神经节和脊髓背角内的表达变化。本文还就dendrin的生物学特征以及在神经系统内表达情况对其近期研究成果和未来发展趋势进行综述。     

Organization of Synaptic Inputs to Rubro-Spinal Neurons in the Cat: Interaction of the Effects of Stimulation of the Cerebellar Nucl. Interpositus and Thalamic Nucl. Ventrolateralis

We studied the interaction of synaptic effects in efferent (rubro-spinal) neurons (RSN) of the red nucleus (NR) of the cat brain using intracellular recording and different variants of stimulation of the cerebellar nucl. interpositus (NI) and nucl. ventrolateralis (VL) of the thalamus. Combined simultaneous or consequent stimulation of the above two inputs allowed us to use collision of the impulses in pre-synaptic pathways for estimation of the peculiarities of summation of post-synaptic potentials; it was taken into consideration that cerebello-rubral fibers are collaterals of the cerebello-thalamic axons. In the case of relatively low intensities of stimulation of the VL and NI, EPSP evoked by these stimulations in RSN could demonstrate linear summation, which is indicative of the absence of common fibers excited by stimulation of both the thalamic and cerebellar structures and of the absence of interaction of synaptic effects on the membrane of the postsynaptic unit (an RSN). With increased stimulation intensity and definite time relations between the stimuli applied to the above structures, the second EPSP evoked by the two stimuli was significantly suppressed; this resulted from a decrease of the afferent drive coming to the NR due to collision of afferent impulses in presynaptic pathways. When an interstimulus interval was increased, the second EPSP could be facilitated (probably due to generation of repetitive impulses in common presynaptic fibers). Our experiments show that the statement concerning the nature of cerebello-rubral inputs (all these inputs are formed by collaterals of the cerebello-(interposito)-thalamic pathways) should be considered with reservation.     

Predominant Neuritic Pathology Induced by Viral Overexpression of <Emphasis Type="Bold">α</Emphasis>-Synuclein in Cell Culture

1. Alpha-synuclein is known to play an important role in the pathogenesis of Parkinson’s disease (PD). The pathogenicity of α-synuclein is related to its ability to form intraneuronal inclusions. The inclusions, which are found in brains of patients with PD and diffuse Lewy body disease consist partially of C-terminally truncated α-synuclein. This α-synuclein species has an increased ability to form aggregates compared to full length α-synuclein. 2. We have used an adeno-associated virus (AAV) vector system to overexpress either C-terminally truncated or full length α-synuclein containing the A53T mutation, which have both been identified in brains of familial PD patients and transgenic mouse models. Dissociated mesencephalic neurons, cerebellar granule neurons, and organotypic midbrain slice cultures were infected with AAV containing the transgene under the control of the cytomegalovirus promoter. 3. We demonstrate that viral overexpression of α-synuclein(A53T) leads to the formation of distorted neurites, intraneuritic swellings, and granular perikaryal deposits in cultured neurons. Our results indicate that these cell culture models may represent an early phase of PD reflecting pathologic neuritic alterations before significant neuronal cell loss occurs.     

Tachykinins Potentiate N-Methyl-D-Aspartate Responses in Acutely Isolated Neurons from the Dorsal Horn

Abstract: Substance P and neurokinin A both potentiated N -methyl- d -aspartate (NMDA)-induced currents recorded in acutely isolated neurons from the dorsal horn of the rat. To elucidate the mechanism underlying this phenomenon, we measured the effects of tachykinins and glutamate receptor agonists on [Ca²⁺]_i in these cells. Substance P, but not neurokinin A, increased [Ca²⁺]_i in a subpopulation of neurons. The increase in [Ca²⁺]_i was found to be due to Ca²⁺ influx through voltage-sensitive Ca²⁺ channels. Substance P and neurokinin A also potentiated the increase in [Ca²⁺]_i produced by NMDA, but not by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid, kainate, or 50 m M K⁺. Phorbol esters enhanced the effects of NMDA and staurosporine inhibited the potentiation of NMDA effects by tachykinins. It is concluded that activation of protein kinase C may mediate the enhancement of NMDA effects by tachykinins in these cells. However, the effects of tachykinins on [Ca²⁺]_i can be dissociated from their effects on NMDA receptors.     

Immunocytochemical demonstration of separate vasopressin-neurophysin and oxytocin-neurophysin neurons in the human hypothalamus

Summary With the use of immunocytochemistry, it was shown that both the supraoptic and paraventricular hypothalamic nuclei in humans contain at least two different neurophysins. These two human neurophysins are immunologically related to bovine neurophysin I and neurophysin II, respectively. One human neurophysin is associated with vasopressin, the other with oxytocin. Human vasopressin-neurophysin and oxytocin-neurophysin are located separately in two different types of neurons, which correspond respectively to the vasopressinergic and oxytocinergic neurons of both the supraoptic and paraventricular nuclei. The neurophysin of the human vasopressinergic suprachiasmatic neurons appears to be closely related to or identical with neurophysin of the vasopressinergic neurons of the human magnocellular hypothalamic nuclei.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

An In-vitro Preparation of Isolated Enteric Neurons and Glia from the Myenteric Plexus of the Adult Mouse

The enteric nervous system is a vast network of neurons and glia running the length of the gastrointestinal tract that functionally controls gastrointestinal motility. A procedure for the isolation and culture of a mixed population of neurons and glia from the myenteric plexus is described. The primary cultures can be maintained for over 7 days, with connections developing among the neurons and glia. The longitudinal muscle strip with the attached myenteric plexus is stripped from the underlying circular muscle of the mouse ileum or colon and subjected to enzymatic digestion. In sterile conditions, the isolated neuronal and glia population are preserved within the pellet following centrifugation and plated on coverslips. Within 24-48 hr, neurite outgrowth occurs and neurons can be identified by pan-neuronal markers. After two days in culture, isolated neurons fire action potentials as observed by patch clamp studies. Furthermore, enteric glia can also be identified by GFAP staining. A network of neurons and glia in close apposition forms within 5 - 7 days. Enteric neurons can be individually and directly studied using methods such as immunohistochemistry, electrophysiology, calcium imaging, and single-cell PCR. Furthermore, this procedure can be performed in genetically modified animals. This methodology is simple to perform and inexpensive. Overall, this protocol exposes the components of the enteric nervous system in an easily manipulated manner so that we may better discover the functionality of the ENS in normal and disease states.