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By comparing behavioral performance and cortical theta activity (4-8 Hz) on a trial by trial basis we examined how the different behavioral stages of tone-induced avoidance learning in the shuttle-box may be distinguishable by theta power as a potential correlate of changing strategies of information processing. Electrocorticograms with pronounced theta content were recorded across the cortical surface of gerbils during avoidance learning and analyzed in each trial in conjunction with reaction times and unconditioned and conditioned responses. The focus of theta analysis in this paradigm with a 5-s delay between tone and foot-shock onsets was on the 14-s periods after hurdle crossing where feedback information from a trial is available. The strongest theta activity occurred in stage 1 of initial tone conditioning which was sharply reduced to a minimum during stage 2 of optimization of unconditioned escape responses from the foot shock. A few initial successful avoidance responses gave rise to a reversal of the decline of theta activity that later reached a second maximum. A systematic increase of theta activity during this stage 3 of avoidance conditioning was found for the occasional trials with unconditioned responses and not for the increasing number of conditioned responses suggesting that error processing is a major correlate of this new increase of theta power. After the second maximum the theta power slowly declined together with a further improvement of behavioral performance indicating that stage 4 of retrieval of the consolidated avoidance response was reached. The results suggest that behind a previously reported general trend of decreasing theta power with increasing performance in this paradigm there is a hidden microstructure of theta activity across trials which separates stages of avoidance conditioning and is partially mirrored by known changes of prefrontal dopamine release.

Recently, we cloned and began to characterize a new N-methyl-D-aspartate receptor (NMDAR) subunit, NR3A. Here we extend our earlier findings by showing that recombinantly expressed NR3A in COS cells is biochemically associated with both NR1 and NR2 subunits. In the oocyte or HEK 293 cell expression systems, co-injection of NR3A with NR1/NR2 subunits acts in a dominant-interfering manner, resulting in a decrease in NMDAR unitary conductance, decrease in Ca(2+) permeability, decrease in Mg(2+) sensitivity, and slight increase in mean open time compared with NR1/NR2 channels. The smaller unitary conductance channel has also been observed in primary cortical neurons cultured from wild-type rodent on postnatal day 8 (P8) and similarly found to be relatively insensitive to Mg(2+) block. Consistent with these findings, whole cell NMDA-evoked currents are larger in NR3A-deficient mice compared with wild-type mice, and this effect follows a developmental pattern similar to that of NR3A protein expression on Western blots, with peak expression at P8. Finally, a new longer splice variant of NR3A has been cloned and found to be expressed in rodent cortical neurons by single-cell RT-PCR and in situ hybridization.

The NMDA (N-methyl-D-aspartate) subclass of glutamate receptor is essential for the synaptic plasticity thought to underlie learning and memory and for synaptic refinement during development. It is currently believed that the NMDA receptor (NMDAR) is a heteromultimeric channel comprising the ubiquitous NR1 subunit and at least one regionally localized NR2 subunit. Here we report the characterization of a regulatory NMDAR subunit, NR3A (formerly termed NMDAR-L or chi-1), which is expressed primarily during brain development. NR3A co-immunoprecipitates with receptor subunits NR1 and NR2 in cerebrocortical extracts. In single-channel recordings from Xenopus oocytes, addition of NR3A to NR1 and NR2 leads to the appearance of a smaller unitary conductance. Genetic knockout of NR3A in mice results in enhanced NMDA responses and increased dendritic spines in early postnatal cerebrocortical neurons. These data suggest that NR3A is involved in the development of synaptic elements by modulating NMDAR activity.

Mice of the inbred strains C57BL/6 and DBA/2 show strain-dependent behavioural differences which have been correlated with variations in brain cholinergic systems. In the present study, the density of muscarinic cholinergic receptors in both strains of mice was determined by autoradiographic methods using [3H]quinuclidinyl benzilate (QNB) and [3H]pirenzepine as ligands. C57BL/6 mice showed a significantly lower [3H]QNB binding level in the frontal cortex by one third as compared to DBA/2 mice. In the striatum and the cholinergic pontomesencephalic nucleus laterodorsalis tegmenti the [3H]QNB binding was lower in C57BL/6 by 28% and 31%, respectively. The [3H]pirenzepine binding level was found to be significantly higher in C57BL/6 temporal cortex (by 22%). These results are discussed in relation to interstrain differences in cholinergic cell density and in the activity of cholinergic enzymes.

A method for the rapid dissection of circumscribed areas of rat cortex is described. The technique does not depend on skull-derived landmarks but uses for stereotaxic orientation the cross-point of the interhemispheric gap with the caudal margin of the cortex. An application of this dissection method to the biochemical analysis of cholinergic markers within the hindlimb representation of the primary somatosensory cortex revealed that both the activity of the enzyme choline acetyltransferase as well as the binding of [3H] quinuclidinyl benzilate to muscarinic cholinergic receptors do not seem to be affected drastically three days after unilateral transection of the sciatic nerve. The only significant effect detected was a slight decrease in the activity of the choline acetyltransferase within the hindlimb representation of the primary somatosensory cortex contralateral to the transected sciatic nerve. In the primary visual cortex, the cholinergic markers investigated did not show significant alterations after sciatic nerve injury.

The efficiency of separation of the radioligand receptor complex from the unbound ligand by both rapid filtration and centrifugation is compared for two standard receptor assays in the rat brain: the beta-adrenergic receptor using 3H-dihydroalprenolol and the benzodiazepine receptor using 3H-flunitrazepam. Whereas in beta-adrenergic binding studies both separation procedures yield similar binding data, the apparent binding in the benzodiazepine receptor assay is lower in the filtration experiments as compared to the data obtained by centrifugation, indicating an incomplete retention of the ligand receptor complex by filter. However, in both assays the results of the binding studies are highly dependent on the type of filters used. The following filters were compared: glass fibre filters GE/B and GF/C (Whatman Ltd., Kent, England), membrane filter HAWP (pore size 0.45 micrometer, Millipore Corporation, Bedford, U.S.A.) and glass fibre filters NK I, NK VIII, NK-15-90, NK 13-90, NK 14-120 (all produced by VEB Feinpapierfabrik Neu-Kaliss, G.D.R.). Although a somewhat higher specific binding can be obtained by the membrane filter and several types of the NK filters as compared to the most widely used Whatman GF/C the manifold higher unspecific binding of the radioligand by these filters does not favour their use in ligand assays. Retention as well as unspecific binding of the filter NK VIII is very similar to the Whatman GF/C filter. This new filter seems, therefore, well suited for radioligand studies.

The postnatal development of [3H]flunitrazepam binding to benzodiazepine receptors has been studied in the retina, lateral geniculate nucleus, superior colliculus, frontal cortex and visual cortex of the rat. In the frontal and visual cortices, binding reached the highest level at postnatal day 25 and then decreased slightly until adulthood. In the superior colliculus, the adult value of [3H]flunitrazepam binding is already reached at postnatal day 25, whereas in the retina the highest binding was found at postnatal day 50. The ontogeny of benzodiazepine binding sites in the visual regions does not essentially differ from that in other brain regions, suggesting that the appearance of [3H]flunitrazepam binding sites in the visual system is not correlated with the development of retinal function and the functional maturation of the visual system with regard to processing of light stimuli. Raising rats in complete darkness from birth until the age of 25 days resulted in significantly decreased binding levels in the lateral geniculate nucleus and in the superior colliculus by 29% and 17%, respectively, as compared to controls. [3H]Flunitrazepam binding in the other regions studied was not affected by dark-rearing. Presumably, the complete lack of visual experience interferes with the functional development of GABAergic mechanisms involved in the gating function of the subcortical visual centres for visual information transfer.