Search

Cortical neurons in brain slices display intrinsic spike frequency adaptation (I-SFA) to constant current inputs, while extracellular recordings show extrinsic SFA (E-SFA) during sustained visual stimulation. Inferring how I-SFA contributes to E-SFA during behavior is challenging due to the isolated nature of slice recordings. To address this, we recorded macaque lateral prefrontal cortex (LPFC) neurons in vivo during a visually guided saccade task and in vitro in brain slices. Broad-spiking (BS) putative pyramidal cells and narrow-spiking (NS) putative inhibitory interneurons exhibit both E-SFA and I-SFA. Developing a data-driven hybrid circuit model comprising NS model neurons receiving BS input reveals that NS model neurons exhibit longer SFA than observed in vivo; however, adding feedforward inhibition corrects this in a manner dependent on I-SFA. Identification of this circuit motif shaping E-SFA in LPFC highlights the roles of both intrinsic and network mechanisms in neural activity underlying behavior., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Can the transcriptomic profile of a neuron predict its physiological properties? Using a Patch-seq dataset of the primary visual cortex, we addressed this question by focusing on spike rate adaptation (SRA), a well-known phenomenon that depends on small conductance calcium (Ca)-dependent potassium (SK) channels. We first show that in parvalbumin-expressing (PV) and somatostatin-expressing (SST) interneurons (INs), expression levels of genes encoding the ion channels underlying action potential generation are correlated with the half-width (HW) of spikes. Surprisingly, the SK encoding gene is not correlated with the degree of SRA (dAdap). Instead, genes that encode proteins upstream from the SK current are correlated with dAdap, a finding validated by a different dataset from the mouse's primary motor cortex that includes pyramidal cells and interneurons, as well as physiological datasets from multiple regions of macaque monkeys. Finally, we construct a minimal model to reproduce observed heterogeneity across cells, with testable predictions.

Mouse primary somatosensory barrel cortex (wS1) processes whisker sensory information, receiving input from two distinct thalamic nuclei. The first-order ventral posterior medial (VPM) somatosensory thalamic nucleus most densely innervates layer 4 (L4) barrels, whereas the higher-order posterior thalamic nucleus (medial part, POm) most densely innervates L1 and L5A. We optogenetically stimulated VPM or POm axons, and recorded evoked excitatory postsynaptic potentials (EPSPs) in different cell-types across cortical layers in wS1. We found that excitatory neurons and parvalbumin-expressing inhibitory neurons received the largest EPSPs, dominated by VPM input to L4 and POm input to L5A. In contrast, somatostatin-expressing inhibitory neurons received very little input from either pathway in any layer. Vasoactive intestinal peptide-expressing inhibitory neurons received an intermediate level of excitatory input with less apparent layer-specificity. Our data help understand how wS1 neocortical microcircuits might process and integrate sensory and higher-order inputs., Competing Interests: BS, PT, MF, JM, TO, OY, JS, CP No competing interests declared, (© 2019, Sermet et al.)

Results of laboratory batch studies often differ from those of outdoor lysimeter or field plot experiments--with respect to degradation as well as sorption. Laboratory micro-lysimeters are a useful device for closing the gap between laboratory and field by both including relevant transport processes in undisturbed soil columns and allowing controlled boundary conditions. In this study, sorption and degradation of the herbicide metsulfuron-methyl in a loamy silt soil were investigated by applying inverse modelling techniques to data sets from different experimental approaches under laboratory conditions at a temperature of 10 degrees C: first, batch-degradation studies and, second, column experiments with undisturbed soil cores (28 cm length x 21 cm diameter). The column experiments included leachate and soil profile analysis at two different run times. A sequential extraction method was applied in both study parts in order to determine different binding states of the test item within the soil. Data were modelled using ModelMaker and Hydrus-1D/2D. Metsulfuron-methyl half-life in the batch-experiments (t1/2 = 66 days) was shown to be about four times higher than in the micro-lysimeter studies (t1/2 about 17 days). Kinetic sorption was found to be a significant process both in batch and column experiments. Applying the one-rate-two-site kinetic sorption model to the sequential extraction data, it was possible to associate the stronger bonded fraction of metsulfuron-methyl with its kinetically sorbed fraction in the model. Although the columns exhibited strong significance of multi-domain flow (soil heterogeneity), the comparison between bromide and metsulfuron-methyl leaching and profile data showed clear evidence for kinetic sorption effects. The use of soil profile data had significant impact on parameter estimates concerning sorption and degradation. The simulated leaching of metsulfuron-methyl as it resulted from parameter estimation was shown to decrease when soil profile data were considered in the parameter estimation procedure. Moreover, it was shown that the significance of kinetic sorption can only be demonstrated by the additional use of soil profile data in parameter estimation. Thus, the exclusive use of efflux data from leaching experiments at any scale can lead to fundamental misunderstandings of the underlying processes.

Using the novel radioligand, [(3)H]-9'-nor-fusicoccin-8'-alcohol, high affinity binding sites for fusicoccin were characterized in preparations from leaves of Arabidopsis thaliana (L.) Heynh. The binding site copartitioned with the plasmalemma marker, vanadate-sensitive K(+), Mg(2+)-ATPase, when microsomal fractions were further purified by aqueous two-phase partitioning in polyethylene glycol-dextran phase systems and sedimented at an equilibrium density of 1.17 grams per cubic centimeter in continuous sucrose density gradients, as did the ATPase marker. The binding of [(3)H]-9'-nor-fusicoccin-8'-alcohol was saturable and Scatchard analysis revealed a biphasic plot with two apparent dissociation constants (K(D)), K(D1) = 1.5 nanomolar and K(D2) = 42 nanomolar, for the radioligand. Binding was optimal at pH 6, thermolabile, and was reduced by 70% when the membrane vesicles were pretreated with trypsin. The data are consistent with the presence of one or several binding proteins for fusicoccin at the plasma membrane of A. thaliana. Binding of the radioligand was unaffected by pretreatment of the sites with various alkylating and reducing agents, but was reduced by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, diethylpyrocarbonate, chloramine T, and periodate. A number of detergents were tested to find optimum conditions for solubilization. Nonanoyl-N-methylglucamide (50 millimolar) solubilized 70% of the radioligand-binding protein complex in undissociated form. Photoaffinity labeling of membrane preparations with a tritiated azido analog of fusicoccin resulted in the labeling of a 34 +/- 1 kilodalton polypeptide. Labeling of this polypeptide, presumably the fusicoccin-binding protein, was severely reduced in the presence of unlabeled fusicoccin.

Monoclonal antibodies were raised against fusicoccin. The toxin, linked to bovine serum albumin through its t-pentenyl moiety, served as immunogen. Hybridomas secreting anti-fusicoccin antibodies were screened by radioimmunoassay employing a novel radioactive derivative, [(3)H]-nor-fusicoccin-alcohol of high specific activity (1.5 x 10(14)Bq/mole). The two monoclonal antibodies reported here are of high apparent affinity for fusicoccin (0.71 x 10(-9) molar and 1.85 x 10(-9) molar). This is comparable to the apparent affinity of rabbit antiserum raised against the same type of conjugate (9.3 x 10(-9) molar). A method for the single step purification of the monoclonal antibodies from ascites fluid is reported. A solid-phase immunoassay, using alkaline phosphatase as enzyme, exhibits a measuring range from 0.1 to 1.5 picomoles (about 70 picograms to 1 nanogram) of fusicoccin. The displacement of [(3)H]-nor-fusicoccin-alcohol from the antibodies by compounds structurally related to fusicoccin exhibits similar selectivity as a microsomal binding assay with the same tracer as radiolabeled probe.

The high-affinity fusicoccin-binding protein (FCBP) was solubilized from plasma-membrane vesicles prepared from leaves of Vicia faba L. by aqueous two-phase partitioning. Conditions for the solubilization of intact FCBP-radioligand complexes were worked out. About 60-70% of the complexes can be solubilized with 50-60 mM nonanoyl-N-methylglucamide in the presence of 1 mg· ml(-1) soybean phosphatidylcholine, type IV S, and 20% (v/v) glycerol at pH 5.5. The slow dissociation of the radioligand, 9'-nor-fusicoccin-8'-alcohol-[(3)H] from the FCBP at low temperatures permits the purification of FCBP-radioligand complexes at 4-10° C by fast protein liquid chromatography on anion-exchange and gel permeation columns. The FCBP, extracted from plasma membranes with cholate and chromatographed in the presence of this detergent, gave an apparent molecular mass (Mr) of 80±20 kDa on gel permeation columns under the conditions used. By comparison of the elution profiles of the fraction most enriched in FCBP-radioligand complexes with polypeptide patterns obtained on sodium dodecyl sulfate-polyacrylamide gels, a polypeptide with an Mr of approx. 34kDa co-separated with the radioactivity profile. A second, faint band of approx. 31 kDa was sometimes also observed co-electrophoresing. Photoaffinity labeling of plasma-membrane vesicles with the new compound 9'-nor-8'[(3,5-[(3)H]-4-azidobenzoy)ethylenediamine]-fusicoccin ([(3)H]ABE-FC) and subsequent separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis labeled a single band with an Mr of 35±1 kDa. Labeling in this band was strongly reduced when the membranes were incubated with [(3)H]ABE-FC in the presence of 0.1-1 μM fusicoccin. From our data, we conclude (i) that the 34-35-kDa polypeptide represents the FCBP and (ii) that in detergent extracts of plasma membranes this polypeptide is probably present as a di- or trimeric structure.