Human Purkinje cells outperform mouse Purkinje cells in dendritic complexity and computational capacity.

Purkinje cells in the cerebellum are among the largest neurons in the brain and have been extensively investigated in rodents. However, their morphological and physiological properties remain poorly understood in humans. In this study, we utilized high-resolution morphological reconstructions and unique electrophysiological recordings of human Purkinje cells ex vivo to generate computational models and estimate computational capacity. An inter-species comparison showed that human Purkinje cell had similar fractal structures but were larger than those of mouse Purkinje cells. Consequently, given a similar spine density (2/µm), human Purkinje cell hosted approximately 7.5 times more dendritic spines than those of mice. Moreover, human Purkinje cells had a higher dendritic complexity than mouse Purkinje cells and usually emitted 2-3 main dendritic trunks instead of one. Intrinsic electro-responsiveness was similar between the two species, but model simulations revealed that the dendrites could process ~6.5 times (n = 51 vs. n = 8) more input patterns in human Purkinje cells than in mouse Purkinje cells. Thus, while human Purkinje cells maintained spike discharge properties similar to those of rodents during evolution, they developed more complex dendrites, enhancing computational capacity.

Stellate cell computational modeling predicts signal filtering in the molecular layer circuit of cerebellum.

The functional properties of cerebellar stellate cells and the way they regulate molecular layer activity are still unclear. We have measured stellate cells electroresponsiveness and their activation by parallel fiber bursts. Stellate cells showed intrinsic pacemaking, along with characteristic responses to depolarization and hyperpolarization, and showed a marked short-term facilitation during repetitive parallel fiber transmission. Spikes were emitted after a lag and only at high frequency, making stellate cells to operate as delay-high-pass filters. A detailed computational model summarizing these physiological properties allowed to explore different functional configurations of the parallel fiber-stellate cell-Purkinje cell circuit. Simulations showed that, following parallel fiber stimulation, Purkinje cells almost linearly increased their response with input frequency, but such an increase was inhibited by stellate cells, which leveled the Purkinje cell gain curve to its 4 Hz value. When reciprocal inhibitory connections between stellate cells were activated, the control of stellate cells over Purkinje cell discharge was maintained only at very high frequencies. These simulations thus predict a new role for stellate cells, which could endow the molecular layer with low-pass and band-pass filtering properties regulating Purkinje cell gain and, along with this, also burst delay and the burst-pause responses pattern.

GSK3 and ß-catenin determines functional expression of sodium channels at the axon initial segment.

Neuronal action potentials are generated through voltage-gated sodium channels, which are tethered by ankyrinG at the membrane of the axon initial segment (AIS). Despite the importance of the AIS in the control of neuronal excitability, the cellular and molecular mechanisms regulating sodium channel expression at the AIS remain elusive. Our results show that GSK3α/ß and ß-catenin phosphorylated by GSK3 (S33/37/T41) are localized at the AIS and are new components of this essential neuronal domain. Pharmacological inhibition of GSK3 or ß-catenin knockdown with shRNAs decreased the levels of phosphorylated-ß-catenin, ankyrinG, and voltage-gated sodium channels at the AIS, both "in vitro" and "in vivo", therefore diminishing neuronal excitability as evaluated via sodium current amplitude and action potential number. Thus, our results suggest a mechanism for the modulation of neuronal excitability through the control of sodium channel density by GSK3 and ß-catenin at the AIS.

Developmental expression of Kv potassium channels at the axon initial segment of cultured hippocampal neurons.

Axonal outgrowth and the formation of the axon initial segment (AIS) are early events in the acquisition of neuronal polarity. The AIS is characterized by a high concentration of voltage-dependent sodium and potassium channels. However, the specific ion channel subunits present and their precise localization in this axonal subdomain vary both during development and among the types of neurons, probably determining their firing characteristics in response to stimulation. Here, we characterize the developmental expression of different subfamilies of voltage-gated potassium channels in the AISs of cultured mouse hippocampal neurons, including subunits Kv1.2, Kv2.2 and Kv7.2. In contrast to the early appearance of voltage-gated sodium channels and the Kv7.2 subunit at the AIS, Kv1.2 and Kv2.2 subunits were tethered at the AIS only after 10 days in vitro. Interestingly, we observed different patterns of Kv1.2 and Kv2.2 subunit expression, with each confined to distinct neuronal populations. The accumulation of Kv1.2 and Kv2.2 subunits at the AIS was dependent on ankyrin G tethering, it was not affected by disruption of the actin cytoskeleton and it was resistant to detergent extraction, as described previously for other AIS proteins. This distribution of potassium channels in the AIS further emphasizes the heterogeneity of this structure in different neuronal populations, as proposed previously, and suggests corresponding differences in action potential regulation.

Colocalization of α-actinin and synaptopodin in the pyramidal cell axon initial segment.

The cisternal organelle that resides in the axon initial segment (AIS) of neocortical and hippocampal pyramidal cells is thought to be involved in regulating the Ca(2+) available to maintain AIS scaffolding proteins, thereby preserving normal AIS structure and function. Through immunocytochemistry and correlative light and electron microscopy, we show here that the actin-binding protein α-actinin is present in the typical cistenal organelle of rodent pyramidal neurons as well as in a large structure in the AIS of a subpopulation of layer V pyramidal cells that we have called the "giant saccular organelle." Indeed, this localization of α-actinin in the AIS is dependent on the integrity of the actin cytoskeleton. Moreover, in the cisternal organelle of cultured hippocampal neurons, α-actinin colocalizes extensively with synaptopodin, a protein that interacts with both actin and α-actinin, and they appear concomitantly during the development of these neurons. Together, these results indicate that α-actinin and the actin cytoskeleton are important components of the cisternal organelle that are probably required to stabilize the AIS.

In vitro maturation of the cisternal organelle in the hippocampal neuron's axon initial segment.

Regulation of Ca(2+) concentrations is essential to maintain the structure and function of the axon initial segment (AIS). The so-called cisternal organelle of the AIS is a structure involved in this regulation, although little is known as to how this organelle matures and is stabilized. Here we describe how the cisternal organelle develops in cultured hippocampal neurons and the interactions that facilitate its stabilization in the AIS. We also characterize the developmental expression of molecules involved in Ca(2+) regulation in the AIS. Our results indicate that synaptopodin (synpo) positive elements considered to be associated to the cisternal organelle are present in the AIS after six days in vitro. There are largely overlapping microdomains containing the inositol 1,4,5-triphosphate receptor 1 (IP(3)R1) and the Ca(2+) binding protein annexin 6, suggesting that the regulation of Ca(2+) concentrations in the AIS is sensitive to IP(3) and subject to regulation by annexin 6. The expression of synpo, IP(3)R1 and annexin 6 in the AIS is independent of the neuron activity, as it was unaffected by tetrodotoxin blockage of action potentials and it was resistant to detergent extraction, indicating that these proteins interact with scaffolding and/or cytoskeleton proteins. The presence of ankyrin G seems to be required for the acquisition and maintenance of the cisternal organelle, while the integrity of the actin cytoskeleton must be maintained for the expression IP(3)R1 and annexin 6 to persist in the AIS.

Casein kinase 2 and microtubules control axon initial segment formation.

The axon initial segment (AIS) is a unique axonal subdomain responsible for the generation of the neuronal action potential and the maintenance of the axon-dendritic functional polarity. Despite its importance, the mechanisms controlling AIS development and maintenance remain largely unknown. Here we show that the AIS microtubule cytoskeleton is composed of a pool of more stable, detergent resistant, microtubules. This AIS specific characteristic is conferred by the presence of CK2, an important regulator of microtubule stability, in the AIS during its development and maturation. We show that CK2α and CK2α' subunits concentrate at the AIS from its initial development, at the same time as pIκBα and ankyrinG. CK2 pharmacological inhibition or suppression of CK2α expression with nucleofected interference RNAs modifies microtubule characteristics throughout the neuron, changes KIF5C distribution, and impairs its own concentration at the AIS, as well as that of ankyrinG, ankyrinG-GFP, pIκBα and voltage gated sodium channels. Moreover, CK2α concentration at the AIS depends on IκBα phosphorylation by IKK and ankyrinG. In conclusion, our results demonstrate a mutual dependence of CK2, ankyrinG and pIκBα for their concentration at the axon initial segment, which is related to the specific characteristics of microtubules at the AIS.

Elementos que influyen en el control metabólico en docentes y administrativos de la UNAN - MANAGUA con Diabetes Mellitus Tipo 2, Managua. Julio - Septiembre 2008

Estudio descriptivo de corte transversal para determinar los elementos que influyen en el control metabólico en docentes y administrativos de la UNAN Managua en las edades comprendidas de 35 a 64 años

New role of IKK alpha/beta phosphorylated I kappa B alpha in axon outgrowth and axon initial segment development.

Neuronal polarity development begins by the outgrowth of the axon and the formation of the axon initial segment which acts as a diffusion barrier and it is the place of action potential generation. The mechanisms controlling this development are largely unknown. We describe a role for IkappaB alpha, the NFkappaB inhibitor, in the initial stages of axon outgrowth and the development of the axon initial segment. In cultured hippocampal neurons, inhibition of IkappaB alpha phosphorylation by IkappaB kinases (IKKs) impedes axon outgrowth. Moreover, the absence of IkappaB alpha phosphorylation, in the next stages of axon development, impairs the localization of structural and functional proteins at the axon initial segment, such as ankyrin G and voltage gated sodium channels. These results demonstrate a new role for proteins of the NFkappaB pathway in the acquisition of neuronal polarity and its involvement in the development of the axon initial segment.