Receptor protein tyrosine phosphatase sigma regulates synapse structure, function and plasticity.

The mechanisms that regulate synapse formation and maintenance are incompletely understood. In particular, relatively few inhibitors of synapse formation have been identified. Receptor protein tyrosine phosphatase σ (RPTPσ), a transmembrane tyrosine phosphatase, is widely expressed by neurons in developing and mature mammalian brain, and functions as a receptor for chondroitin sulfate proteoglycans that inhibits axon regeneration following injury. In this study, we address RPTPσ function in the mature brain. We demonstrate increased axon collateral branching in the hippocampus of RPTPσ null mice during normal aging or following chemically induced seizure, indicating that RPTPσ maintains neural circuitry by inhibiting axonal branching. Previous studies demonstrated a role for pre-synaptic RPTPσ promoting synaptic differentiation during development; however, subcellular fractionation revealed enrichment of RPTPσ in post-synaptic densities. We report that neurons lacking RPTPσ have an increased density of pre-synaptic varicosities in vitro and increased dendritic spine density and length in vivo. RPTPσ knockouts exhibit an increased frequency of miniature excitatory post-synaptic currents, and greater paired-pulse facilitation, consistent with increased synapse density but reduced synaptic efficiency. Furthermore, RPTPσ nulls exhibit reduced long-term potentiation and enhanced novel object recognition memory. We conclude that RPTPσ limits synapse number and regulates synapse structure and function in the mature CNS.

Morphological and electrophysiological characteristics of layer V neurons of the rat lateral entorhinal cortex.

The intrinsic electrophysiological and morphological properties of lateral entorhinal area (LEA) layer V neurons were investigated by sharp electrode intracellular recording and biocytin labeling in vitro. The morphological analysis revealed that layer V of the LEA contains three distinct subtypes of principal neurons, which were classified as pyramidal, horizontal, and polymorphic neurons. Pyramidal cells were the most abundant subtype (57%) and could be further subdivided into neurons with large, small, and star-like somas. Similarly to pyramidal cells, horizontal neurons (11%) had a prominent apical dendrite. However, their distinctive basal dendritic plexus extended primarily in the horizontal plane. Polymorphic neurons (32%) were characterized by a multipolar dendritic organization. Electrophysiological analysis of neurons in the three categories demonstrated a diversity of electrophysiological profiles within each category and no significant differences between groups. Neurons in the three subgroups could display instantaneous and/or time-dependent inward rectification and different degrees of spike frequency adaptation. None of the recorded cells displayed an intrinsic oscillatory bursting discharge. Many neurons in the three subgroups, however, displayed slow (3.5-14 Hz), sustained, subthreshold membrane potential oscillations. The morphological and electrophysiological diversity of principal neurons in the LEA parallels that previously reported for the medial entorhinal area and suggests that, with respect to the deep layers, similar information processing is performed across the mediolateral extent of the entorhinal cortex. Layer V of the entorhinal cortex may undertake very complex operations beyond acting as a relay station of hippocampal processed information to the neocortex.

Visualization of the dendritic arbor of neurons in intact 500 microm thick brain slices.

Characterizing the structure and electrophysiological properties of single neurons is essential for understanding how individual cells contribute to the function of neuronal networks. Following intra-cellular recording from neurons in acute brain slices, the structure of the recorded cell has typically been examined by serial sectioning of the tissue slice and then reconstructing the neuron of interest; a labor-intensive and time-consuming process. Here, we have adapted a whole-mount immunohistochemical technique and used it to visualize the dendritic arbor of individual neurons in sections of adult CNS tissue up to 500 microm thick. Permeabilization of the slice and extensive washing allow histochemical reagents to penetrate and be washed from the section, producing limited background staining. Using this method, the cell within the slice can be sectioned optically and reconstructed using the optical sections. We present images of the dendritic trees of neurons in 500 microm thick slices of adult rat entorhinal cortex and hippocampus, labeled either immunohistochemically, or by biocytin injection following whole-cell patch clamp or sharp electrode recordings. The resolution obtained is sufficient to visualize dendritic spines deep within the section. The method is free from artifacts associated with cutting serial sections and is broadly applicable to tasks that require visualization of the fine structure of individual cells in thick slices of CNS tissue.

Cholinergic suppression of excitatory synaptic responses in layer II of the medial entorhinal cortex.

Theta-frequency (4-12 Hz) electroencephalographic activity is thought to play a role in mechanisms mediating sensory and mnemonic processing in the entorhinal cortex and hippocampus, but the effects of acetylcholine on excitatory synaptic inputs to the entorhinal cortex are not well understood. Field excitatory postsynaptic potentials (fEPSPs) evoked by stimulation of the piriform (olfactory) cortex were recorded in the medial entorhinal cortex during behaviors associated with theta activity (active mobility) and were compared with those recorded during nontheta behaviors (awake immobility and slow wave sleep). Synaptic responses were smaller during behavioral activity than during awake immobility and sleep, and responses recorded during movement were largest during the negative phase of the theta rhythm. Systemic administration of cholinergic agonists reduced the amplitude of fEPSPs, and the muscarinic receptor blocker scopolamine strongly enhanced fEPSPs, suggesting that the theta-related suppression of fEPSPs is mediated in part by cholinergic inputs. The reduction in fEPSPs was investigated using in vitro intracellular recordings of EPSPs in Layer II neurons evoked by stimulation of Layer I afferents. Constant bath application of the muscarinic agonist carbachol depolarized membrane potential and suppressed EPSP amplitude in Layer II neurons. The suppression of EPSPs was not associated with a substantial change in input resistance, and could not be accounted for by a depolarization-induced reduction in driving force on the EPSP. The GABA(A) receptor-blocker bicuculline (50 microM) did not prevent the cholinergic suppression of EPSPs, suggesting that the suppression is not dependent on inhibitory mechanisms. Paired-pulse facilitation of field and intracellular EPSPs were enhanced by carbachol, indicating that the suppression is likely due to inhibition of presynaptic glutamate release. These results indicate that, in addition to well known effects on postsynaptic conductances that increase cellular excitability, cholinergic activation in the entorhinal cortex results in a strong reduction in strength of excitatory synaptic inputs from the piriform cortex.

Graded persistent activity in entorhinal cortex neurons.

Working memory represents the ability of the brain to hold externally or internally driven information for relatively short periods of time. Persistent neuronal activity is the elementary process underlying working memory but its cellular basis remains unknown. The most widely accepted hypothesis is that persistent activity is based on synaptic reverberations in recurrent circuits. The entorhinal cortex in the parahippocampal region is crucially involved in the acquisition, consolidation and retrieval of long-term memory traces for which working memory operations are essential. Here we show that individual neurons from layer V of the entorhinal cortex-which link the hippocampus to extensive cortical regions-respond to consecutive stimuli with graded changes in firing frequency that remain stable after each stimulus presentation. In addition, the sustained levels of firing frequency can be either increased or decreased in an input-specific manner. This firing behaviour displays robustness to distractors; it is linked to cholinergic muscarinic receptor activation, and relies on activity-dependent changes of a Ca2+-sensitive cationic current. Such an intrinsic neuronal ability to generate graded persistent activity constitutes an elementary mechanism for working memory.