Michael J. Friedlander

Suppression of PKR Promotes Network Excitability and Enhanced Cognition by Interferon-γ-Mediated Disinhibition

The double-stranded RNA-activated protein kinase (PKR) was originally identified as a sensor of virus infection, but its function in the brain remains unknown. Here, we report that the lack of PKR enhances learning and memory in several behavioral tasks while increasing network excitability. In addition, loss of PKR increases the late phase of long-lasting synaptic potentiation (L-LTP) in hippocampal slices. These effects are caused by an interferon-γ (IFN-γ)-mediated selective reduction in GABAergic synaptic action. Together, our results reveal that PKR finely tunes the network activity that must be maintained while storing a given episode during learning. Because PKR activity is altered in several neurological disorders, this kinase presents a promising new target for the treatment of cognitive dysfunction. As a first step in this direction, we show that a selective PKR inhibitor replicates the Pkr(-/-) phenotype in WT mice, enhancing long-term memory storage and L-LTP.

The kinetic profile of intracellular calcium predicts long-term potentiation and long-term depression.

Efficiency of synaptic transmission within the neocortex is regulated throughout life by experience and activity. Periods of correlated or uncorrelated presynaptic and postsynaptic activity lead to enduring changes in synaptic efficiency [long-term potentiation (LTP) and long-term depression (LTD), respectively]. The initial plasticity triggering event is thought to be a precipitous rise in postsynaptic intracellular calcium, with higher levels inducing LTP and more moderate levels inducing LTD. We used a pairing protocol in visual cortical brain slices from young guinea pigs with whole-cell recording and calcium imaging to compare the kinetic profiles of calcium signals generated in response to individual pairings along with the cumulative calcium wave and plasticity outcome. The identical pairing protocol applied to layer 2/3 pyramidal neurons results in different plasticity outcomes between cells. These differences are not attributable to variations in the conditioning protocol, cellular properties, inter-animal variability, animal age, differences in spike timing between the synaptic response and spikes, washout of plasticity factors, recruitment of inhibition, or activation of different afferents. The different plasticity outcomes are reliably predicted by individual intracellular calcium transients in the dendrites after the first few pairings. In addition to the differences in the individual calcium transients, the cumulative calcium wave that spreads to the soma also has a different profile for cells that undergo LTP versus LTD. We conclude that there are biological differences between like-type cells in the dendritic calcium signals generated by coincident synaptic input and spiking that determine the sign of the plasticity response after brief associations.

Developmental Down-Regulation of LTD in Cortical Layer IV and Its Independence of Modulation by Inhibition

For in vitro LTD to remain viable as a model for synaptic weakening in visual cortical plasticity, it is crucial that it display a critical period for its induction within layer IV. A complicating factor, however, is that LTD in layer IV is modulated by inhibitory postsynaptic potentials (IPSPs); postsynaptic responses characterized as containing IPSPs do not depress in response to 1 Hz afferent stimulation. By blocking IPSPs intracellularly, we find that the ability to induce LTD in layer IV neurons is restored in juvenile, but not in mature animals. This developmental down-regulation of LTD induction is specific for layer IV when compared with LTD induction in layers II/III. These data are consistent with the hypothesis that an LTD-like phenomenon is involved in critical period plasticity and is apparently independent of developmental changes in inhibitory circuitry.

What Can Medical Education Learn From the Neurobiology of Learning

The last several decades have seen a large increase in knowledge of the underlying biological mechanisms that serve learning and memory. The insights gleaned from neurobiological and cognitive neuroscientific experimentation in humans and in animal models have identified many of the processes at the molecular, cellular, and systems levels that occur during learning and the formation, storage, and recall of memories. Moreover, with the advent of noninvasive technologies to monitor patterns of neural activity during various forms of human cognition, the efficacy of different strategies for effective teaching can be compared. Considerable insight has also been developed as to how to most effectively engage these processes to facilitate learning, retention, recall, and effective use and application of the learned information. However, this knowledge has not systematically found its way into the medical education process. Thus, there are considerable opportunities for the integration of current knowledge about the biology of learning with educational strategies and curricular design. By teaching medical students in ways that use this knowledge, there is an opportunity to make medical education easier and more effective. The authors present 10 key aspects of learning that they believe can be incorporated into effective teaching paradigms in multiple ways. They also present recommendations for applying the current knowledge of the neurobiology of learning throughout the medical education continuum.

Structure and projections of white matter neurons in the postnatal rat visual cortex.

Transient contributions of subplate neurons to the initial development of the cortex are well‐characterized, yet little data are available on a subpopulation of subplate neurons that persist in the white matter (WM) of the cerebral cortex across development. To characterize the WM neurons, differential interference contrast and Nomarski optics were used to visualize individual cells in the WM in slices of rat visual cortex at postnatal ages 9–23. Soma‐dendritic morphology and local axonal projection patterns, including probable synaptic innervation sites of their axons, were identified by intracellular filling with biocytin during electrophysiologic recordings. Dendritic branches of all WM neurons, tripartitioned here into upper, middle, and deep divisions, extend throughout the WM and frequently into the overlying cortex. Axonal arborizations from most WM neurons, including apparent boutons, project into adjacent WM with many also innervating overlying cortical layers, whereas some project into the stratum oriens/alveus of the hippocampal formation. Processes of a subset of WM neurons appear to be confined to the WM itself. By using antimicrotubule associated protein (MAP2) immunostaining to quantify the density of WM neurons in rat visual cortex, we find that their overall numbers decrease to approximately 30% of initial levels during postnatal development. During this same developmental period, an increasing percentage of WM neurons contain the synthetic enzyme for nitric oxide, nitric oxide synthase (NOS), as evaluated by immunostaining. Thus, WM neurons that survive the initial perinatal period of cell death are positioned under the laminae of the maturing cortex to potentially modulate the integration of visual signals through either conventional synaptic or nonconventional (diffusible NO signaling) mechanisms. J. Comp. Neurol. 434:233–252, 2001.

Properties of persistent postnatal cortical subplate neurons

Subplate (SP) neurons are important for the proper development of thalamocortical innervation. They are necessary for formation of ocular dominance and orientation columns in visual cortex. During the perinatal period, many SP neurons die. The surviving cohort forms interstitial cells in the white matter (WM) and a band of horizontally oriented cells below layer VI (layer VIb, layer VII, or subplate cells). Although the function of embryonic SP neurons has been well established, the functional roles of WM and postnatal SP cells are not known. We used a combination of anatomical, immunohistochemical, and electrophysiological techniques to explore the dendritic morphology, neurotransmitter phenotype, intrinsic electrophysiological, and synaptic input properties of these surviving cells in the rat visual cortex. The density of SP and WM cells significantly decreases during the first month of life. Both populations express neuronal markers and have extensive dendritic arborizations within the SP, WM, and to the overlying visual cortex. Some intrinsic electrophysiological properties of SP and WM cells are similar: each generates high-frequency slowly adapting trains of action potentials in response to a sustained depolarization. However, SP cells exhibit greater frequency-dependent action potential broadening than WM neurons. Both cell types receive predominantly AMPA/kainate receptor-mediated excitatory synaptic input that undergoes paired-pulse facilitation as well as NMDA receptor and GABAergic input. Synaptic inputs to these cells can also undergo long-term synaptic plasticity. Thus, surviving SP and WM cells are functional electrogenic neurons integrated within the postnatal visual cortical circuit.

Plasticity between Neuronal Pairs in Layer 4 of Visual Cortex Varies with Synapse State

In neocortex, the induction and expression of long-term potentiation (LTP) and long-term depression (LTD) vary depending on cortical area and laminae of presynaptic and postsynaptic neurons. Layer 4 (L4) is the initial site of sensory afference in barrel cortex and primary visual cortex (V1) in which excitatory inputs from thalamus, L6, and neighboring L4 cells are integrated. However, little is known about plasticity within L4. We studied plasticity at excitatory synaptic connections between pairs and triplets of interconnected L4 neurons in guinea pig V1 using a fixed delay pairing protocol. Plasticity outcomes were heterogeneous, with some connections undergoing LTP (n = 7 of 42), some LTD (n = 19 of 42), and some not changing (n = 16 of 42). Although quantal analysis revealed both presynaptic and postsynaptic plasticity expression components, reduction in quantal size (a postsynaptic property) contributing to LTD was ubiquitous, whereas in some cell pairs, this change was overridden by an increase in the probability of neurotransmitter release (a presynaptic property) resulting in LTP. These changes depended on the initial reliability of the connections: highly reliable connections depressed with contributions from presynaptic and postsynaptic effects, and unreliable connections potentiated as a result of the predominance of presynaptic enhancement. Interestingly, very strong, reliable pairs of connected cells showed little plasticity. Pairs of connected cells with a common presynaptic or postsynaptic L4 cell behaved independently, undergoing plasticity of different or opposite signs. Release probability of a connection with initial 100% failure rate was enhanced after pairing, potentially avoiding silencing of the presynaptic terminal and maintaining L4–L4 synapses in a broader dynamic range.

Synaptic Output of Individual Layer 4 Neurons in Guinea Pig Visual Cortex

More than 90% of geniculocortical axons from the dorsal lateral geniculate nucleus of the thalamus innervate layer 4 (L4) of V1 (primary visual cortex). Excitatory neurons, which comprise >80% of the neuronal population in L4, synapse mainly onto adjacent L4 neurons and layer 2/3 (L2/3) neurons. It has been suggested that intralaminar L4–L4 connections contribute to amplifying and refining thalamocortical signals before routing to L2/3. To unambiguously probe the properties of the synaptic outputs from these L4 excitatory neurons, we used multiple simultaneous whole-cell patch-clamp recording and stimulation from two to four neighboring L4 neurons. We recorded uEPSCs (evoked unitary synaptic currents) in response to pairs of action potentials elicited in single presynaptic L4 neurons for 102 L4 cell pairs and found that their properties are more diverse than previously described. Averaged unitary synaptic response peak amplitudes spanned almost three orders of magnitude, from 0.42 to 192.92 pA. Although connections were, on average, reliable (average failure rate, 25%), we recorded a previously unknown subset of unreliable (failure rates from 30 to 100%) and weak (averaged response amplitudes, <5 pA) connections. Reliable connections with high probability of neurotransmitter release responded to paired presynaptic pulses with depression, whereas unreliable connections underwent paired-pulse facilitation. Recordings from interconnected sets of L4 triplets revealed that synaptic response amplitudes and reliability were equally variable between independent cell pairs and those that shared a common presynaptic or postsynaptic cell, suggesting local perisynaptic influences on the development and/or state of synaptic function.

Chapter 19 Temporal covariance of postsynaptic membrane potential and synaptic input – role in synaptic efficacy in visual cortex

Publisher Summary The chapter discusses the role played by temporal covariance of pre- and post-synaptic activity in altering synaptic efficacy. This phenomenon is demonstrated in cells of the primary visual cortex in guinea-pigs and cats, and in more mature animals as well as those at the critical development age for the visual cortex. The results suggest that there is a temporal window about which the pre- and post-synaptic conjunction occurs in order to alter synaptic efficacy. This is particularly important if temporal associations are to be detected and specifically modify the connections that participated in them. The effects occur in the visual cortex beyond the classically defined critical period. The results also suggest that a threshold mechanism operates to only allow the effects of temporal conjunction of pre- and post-synaptic activity to occur outside of a certain “dead zone” around the resting membrane potential. The effects observed appeared to be spatially specific— that is, positive pairing (conjunction of pre-synaptic activation and post-synaptic depolarization) only enhances the efficacy of the paired synapses, and negative pairings (conjunction of pre-synaptic activation and postsynaptic hyper-polarization) only reduce the efficacy of the paired synapses. In some cases, unpaired pathways onto the same cell show no effect, or in other cases, the unpaired pathway shows an opposite effect.

Intracellular injections of permanent tracers in the fixed slice : a comparison of HRP and biocytin

Here we describe a method for intracellularly injecting mixtures of the fluorescent dye Lucifer Yellow and the permanent tracers HRP or biocytin into aldehyde-fixed slices of the dorsal lateral geniculate nucleus in young postnatal cats. Lucifer Yellow was used for visual control in the injection procedure and the inclusion of HRP or biocytin allowed the subsequent use of simple histochemical processing to give a permanent record of the injected cells. Both tracer mixtures revealed the dendritic morphology of injected cells. However, HRP was found to be superior to biocytin, in that dendrites were better defined and fine details of cellular morphology such as spines were consistently revealed. Using this technique we were able to demonstrate that the dendritic morphology of geniculate cells is much more mature between birth and 2 weeks than was thought from previous studies using Golgi methods.