Gianmaria Maccaferri

Social Interaction and Sensorimotor Gating Abnormalities in Mice Lacking Dvl1

Mice completely deficient for Dvl1, one of three mouse homologs of the Drosophila segment polarity gene Dishevelled, were created by gene targeting. Dvl1-deficient mice are viable, fertile, and structurally normal. Surprisingly, these mice exhibited reduced social interaction, including differences in whisker trimming, deficits in nest-building, less huddling contact during home cage sleeping, and subordinate responses in a social dominance test. Sensorimotor gating was abnormal, as measured by deficits in prepulse inhibition of acoustic and tactile startle. Thus, Dvl1 mutants may provide a model for aspects of several human psychiatric disorders. These results are consistent with an interpretation that common genetic mechanisms underlie abnormal social behavior and sensorimotor gating deficits and implicate Dvl1 in processes underlying complex behaviors.

The hyperpolarization‐activated current (Ih) and its contribution to pacemaker activity in rat CA1 hippocampal stratum oriens‐alveus interneurones.

1. The hyperpolarization‐activated current (Ih) and its role in pacemaking activity in rat hippocampal stratum oriens‐alveus interneurones was studied using whole‐cell and perforated patch‐clamp configurations. 2. Voltage‐clamp recordings revealed Ih as a slowly activating, inward current, activated by hyperpolarizing steps (holding potential, Vh = ‐40 mV), with a reversal potential close to ‐30 mV. Its activation curve ranged from approximately ‐50 to ‐120 mV with a mid‐activation point of ‐84.1 mV. 3. Ih was blocked by external application of Cs+ (2‐5 mM) and ZD7288 (100 microM), but not by Ba2+ (1 mM). 4. Ih was potentiated by both noradrenaline and isoprenaline by a mechanism consistent with a shift in the Ih activation curve. 5. Under current‐clamp conditions (Vh = ‐60 mV), ZD7288 induced a membrane hyperpolarization concomitant with an increase in the membrane input resistance and abolished the voltage sag generated by hyperpolarizing current injection. 6. Analysis of the current‐discharge relationship revealed that block of Ih differentially increased the firing frequency of spikes occurring early in the train compared with those occurring late in the discharge. 7. When applied to spontaneously firing cells, ZD7288 reduced the firing frequency by selectively altering the time course of the interspike interval, while minimally affecting other action potential characteristics. Similarly, isoprenaline increased the spontaneous firing frequency by an effect exclusively on the after‐hyperpolarization and interspike interval. 8. These results provide evidence for the involvement of Ih in the excitability and generation of spontaneous firing in hippocampal stratum oriens‐alveus interneurones.

Cell surface domain specific postsynaptic currents evoked by identified GABAergic neurones in rat hippocampus in vitro.

1 Inhibitory postsynaptic currents (IPSCs) evoked in CA1 pyramidal cells (n= 46) by identified interneurones (n= 43) located in str. oriens were recorded in order to compare their functional properties and to determine the effect of synapse location on the apparent IPSC kinetics as recorded using somatic voltage clamp at ‐70 mV and nearly symmetrical [Cl−]. 2 Five types of visualised presynaptic interneurone, oriens‐lacunosum moleculare (O‐LMC), basket (BC), axo‐axonic (AAC), bistratified (BiC) and oriens‐bistratified (O‐BiC) cells, were distinguished by immunocytochemistry and/or synapse location using light and electron microscopy. 3 Somatostatin immunoreactive O‐LMCs, innervating the most distal dendritic shafts and spines, evoked the smallest amplitude (26 ± 10 pA, s.e.m., n= 8) and slowest IPSCs (10‐90 % rise time, 6.2 ± 0.6 ms; decay, 20.8 ± 1.7 ms, n= 8), with no paired‐pulse modulation of the second IPSC (93 ± 4 %) at 100 ms interspike interval. In contrast, parvalbumin‐positive AACs evoked larger amplitude (308 ± 103 pA, n= 7) and kinetically faster (rise time, 0.8 ± 0.1 ms; decay 11.2 ± 0.9 ms, n= 7) IPSCs showing paired‐pulse depression (to 68 ± 5 %, n= 6). Parvalbumin‐ or CCK‐positive BCs (n= 9) terminating on soma/dendrites, BiCs (n= 4) and O‐BiCs (n=7) innervating dendrites evoked IPSCs with intermediate kinetic parameters. The properties of IPSCs and sensitivity to bicuculline indicated that they were mediated by GABAA receptors. 4 In three cases, kinetically complex, multiphasic IPSCs, evoked by an action potential in the recorded basket cells, suggested that coupled interneurones, possibly through electrotonic junctions, converged on the same postsynaptic neurone. 5 The population of O‐BiCs (4 of 4 somatostatin positive) characterised in this study had horizontal dendrites restricted to str. oriens/alveus and innervated stratum radiatum and oriens. Other BiCs had radial dendrites as described earlier. The parameters of IPSCs evoked by BiCs and O‐BiCs showed the largest cell to cell variation, and a single interneurone could evoke both small and slow as well as large and relatively fast IPSCs. 6 The kinetic properties of the somatically recorded postsynaptic current are correlated with the innervated cell surface domain. A significant correlation of rise and decay times for the overall population of unitary IPSCs suggests that electrotonic filtering of distal responses is a major factor for the location and cell type specific differences of unitary IPSCs, but molecular heterogeneity of postsynaptic GABAA receptors may also contribute to the observed kinetic differences. Furthermore, domain specific differences in the short‐term plasticity of the postsynaptic response indicate a differentiation of interneurones in activity‐dependent responses.

Passive propagation of LTD to stratum oriens-alveus inhibitory neurons modulates the temporoammonic input to the hippocampal CA1 region

Excitatory synaptic activity in horizontal stratum oriens-alveus interneurons (OAIs) is driven by the recurrent collaterals of CA1 pyramidal cells and is strongly influenced by protocols that elicit synaptic plasticity in these principal neurons. Induction of LTD in the Schaffer collateral-CA1 pyramidal neuron synapse causes a passive down-regulation of stratum radiatum-evoked excitatory synaptic responses onto OAIs. In addition, we show that the strength of the temporoammonic input to the CA1 pyramidal neuron distal dendrites is regulated by OAI activity. The passive propagation of LTD to OAIs consequently disinhibits the direct entorhinal cortex-CA1 input, resulting in an enhanced excitation of CA1 pyramidal neurons by a mechanism not requiring activation of the trisynaptic pathway.

Interneuron Diversity series: Hippocampal interneuron classifications – making things as simple as possible, not simpler

The nervous system is made up of many specific types of neuron intricately intertwined to form complex networks. Identifying and defining the characteristic features of the many different neuronal types is essential for achieving a cellular understanding of complex activity from perception to cognition. So far, cortical GABAergic interneurons have represented the epitome of cellular diversity in the CNS. Despite the desperate need for effective classification criteria allowing a common language among neuroscientists, interneurons still evoke memories of Babel. Several approaches are now available to overcome the challenges and problems associated with the various classification systems used so far.

Cholecystokinin-immunopositive basket and Schaffer collateral-associated interneurones target different domains of pyramidal cells in the CA1 area of the rat hippocampus.

Two types of GABAergic interneurone are known to express cholecystokinin-related peptides in the isocortex: basket cells, which preferentially innervate the somata and proximal dendrites of pyramidal cells; and double bouquet cells, which innervate distal dendrites and dendritic spines. In the hippocampus, cholecystokinin immunoreactivity has only been reported in basket cells. However, at least eight distinct GABAergic interneurone types terminate in the dendritic domain of CA1 pyramidal cells, some of them with as yet undetermined neurochemical characteristics. In order to establish whether more than one population of cholecystokinin-expressing interneurone exist in the hippocampus, we have performed whole-cell current clamp recordings from interneurones located in the stratum radiatum of the hippocampal CA1 region of developing rats. Recorded neurones were filled with biocytin to reveal their axonal targets, and were tested for the presence of pro-cholecystokinin immunoreactivity. The results show that two populations of cholecystokinin-immunoreactive interneurones exist in the CA1 area (n=15 positive cells). Cholecystokinin-positive basket cells (53%) preferentially innervate stratum pyramidale and adjacent strata oriens and radiatum. A second population of cholecystokinin-positive cells, previously described as Schaffer collateral-associated interneurones [Vida et al. (1998) J. Physiol. 506, 755-773], have axons that ramify almost exclusively in strata radiatum and oriens, overlapping with the Schaffer collateral/commissural pathway originating from CA3 pyramidal cells. Two of seven of the Schaffer collateral-associated cells were also immunopositive for calbindin. Soma position and orientation in stratum radiatum, the number and orientation of dendrites, and the passive and active membrane properties of the two cell populations are only slightly different. In addition, in stratum radiatum and its border with lacunosum of perfusion-fixed hippocampi, 31.6+/-3.8% (adult) or 26.8+/-2.9% (postnatal day 17-20) of cholecystokinin-positive cells were also immunoreactive for calbindin. Therefore, at least two populations of pro-cholecystokinin-immunopositive interneurones, basket and Schaffer collateral-associated cells, exist in the CA1 area of the hippocampus, and are probably homologous to cholecystokinin-immunopositive basket and double bouquet cells in the isocortex. It is not known if the GABAergic terminals of double bouquet cells are co-aligned with specific glutamatergic inputs. However, in the hippocampal CA1 area, it is clear that the terminals of Schaffer collateral-associated cells are co-stratified with the glutamatergic input from the CA3 area, with as yet unknown functional consequences. The division of the postsynaptic neuronal surface by two classes of GABAergic cell expressing cholecystokinin in both the hippocampus and isocortex provides further evidence for the uniform synaptic organisation of the cerebral cortex.

Stratum oriens horizontal interneurone diversity and hippocampal network dynamics

In this last decade, the combination of differential interference contrast infrared video technology and patch‐clamp techniques applied to slices in vitro has allowed the routine electrophysiological recording of visually identified central neurones. This has opened the way to the possibility of preselecting GABAergic interneurones of the hippocampus on the basis of some peculiar morphological characteristics. In particular, stratum oriens ‘horizontal’ interneurones are easily recognizable in living hippocampal slices because of their location and bipolar/bitufted appearance. Thus, this class of cells has rapidly risen as one of the most studied in the entire hippocampus. In this review, I will try to assemble the vast electrophysiological knowledge on these interneurones into a more focused picture, which is relevant for network activity in vitro and in vivo.

Electrical Coupling between Interneurons with Different Excitable Properties in the Stratum Lacunosum-Moleculare of the Juvenile CA1 Rat Hippocampus

Electrical coupling among GABAergic interneurons is believed to play an essential role in shaping synchronized brain network activity related to cognition and behavior. We have studied the rules governing the electrical coupling between hippocampal interneurons located in stratum lacunosum-moleculare of the CA1 hippocampus. The most frequently recorded interneuron subtype had short multipolar dendrites and a dense local axonal arborization, typical of neurogliaform cells. Electrical excitability in this class of interneurons was heterogeneous. Although injection of small current steps evoked late spiking, larger steps triggered different types of firing patterns. Trains of action potentials ranged from clearly adapting to highly irregular, with clustered or mostly regular spikes. Electrotonic and action potentials could be propagated to the coupled cells; the coupling coefficient for electrotonic signals was 0.035, which compared with 0.005 for action potentials. Electrical coupling was reversibly blocked by application of carbenoxolone. Multiple simultaneous recordings indicated that interneurons with similar and different firing patterns were electrically coupled. This visual impression was quantitatively confirmed by principal component analysis applied to variables related to membrane excitability. In fact, the probability of finding electrically coupled neurons in our sample was not dependent on the excitable properties of the cells tested and was ∼0.34. The presence of diffuse electrical coupling among hippocampal interneurons of stratum lacunosum-moleculare with different excitability is a novel finding with important implications. For example, the promiscuity of electrical connections may endow inhibitory networks with a large degree of flexibility and regulate the computational power of the hippocampus during different synchronized states.

Noradrenergic Modulation of Electrical Coupling in GABAergic Networks of the Hippocampus

Noradrenergic modulation of cortical circuits is involved in information processing, regulation of higher functions, and prevention of epileptic activity. Here, we studied the effects of noradrenaline on the functional connectivity of GABAergic networks of the hippocampus and show that electrical synapses between interneurons are a novel target of noradrenergic modulation in vitro. Application of noradrenaline or of the selective β-adrenergic agonist isoproterenol decreased gap junction-based coupling in paired recordings from stratum lacunosum-moleculare interneurons by ∼40%. Similar results were obtained after pharmacological stimulation of the adenylyl cyclase with forskolin. In contrast, the adenylyl cyclase antagonist MDL12330A [cis-N-(2-phenylcyclopentyl)azacyclotridec-1-en-2-amine] or the specific protein kinase A (PKA) inhibitor H89 (N-[2-(p-bromocinnamyl-amino)ethyl]-5-isoquinolinesulfonamide dihydrochloride) enhanced the basal strength of coupling by ∼30%. In addition, PKA-mediated phosphorylation was critical for both isoproterenol- and forskolin-dependent regulation of coupling, because inclusion of the PKA antagonist KT5720 [(9S,10R,12R)-2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3′,2′,1′-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylicacid hexyl ester] in the recording pipettes prevented modulation. Lastly, we studied the effects of β-adrenergic modulation on mixed polysynaptic transmission within the GABAergic network. Isoproterenol depressed propagation of GABAA receptor-mediated synaptic currents, but did not change significantly direct GABAergic input, indicating that regulation of electrical coupling adds flexibility to the information flow generated by chemical synapses. In conclusion, activation of β-adrenergic receptors in stratum lacunosum-moleculare GABAergic networks reduces electrical synaptic transmission via a cAMP/PKA signaling cascade, and affects the degree of synaptic divergence within the circuit. We propose that this dynamic modulation and interplay between electrical and chemical synaptic transmission in GABAergic networks contributes to the tuning of memory processes in vivo, and prevents hypersynchronous activity.

Propagation of postsynaptic currents and potentials via gap junctions in GABAergic networks of the rat hippocampus

The integration of synaptic signalling in the mammalian hippocampus underlies higher cognitive functions such as learning and memory. We have studied the gap junction‐mediated cell‐to‐cell and network propagation of GABAA receptor‐mediated events in stratum lacunosum moleculare interneurons of the rat hippocampus. Propagated events were identified both in voltage‐ and current‐clamp configurations. After blockade of ionotropic excitatory synaptic transmission, voltage‐clamp recordings with chloride‐loaded electrodes (predicted GABAA receptor reversal potential: 0 mV) at −15 mV revealed the unexpected presence of spontaneous events of opposite polarities. Inward events were larger and kinetically faster when compared to outward currents. Both types of events were blocked by gabazine, but only outward currents were significantly affected by the gap junction blocker carbenoxolone, indicating that outward events originated in electrically coupled neurons. These results were in agreement with computational modelling showing that propagated events were modulated in size and shape by their relative distance to the gap junction site. Paired recordings from electrically coupled interneurons performed with high‐ and low‐chloride pipettes (predicted GABAA receptor reversal potentials: 0 mV and −80 mV, respectively) directly demonstrated that depolarizing postsynaptic events could propagate to the cell recorded with the low‐chloride solution. Cell‐to‐cell propagation was abolished by carbenoxolone, and was not observed in uncoupled pairs. Application of 4‐aminopyridine on slices resulted in spontaneous network activation of interneurons, which was driven by excitatory GABAA receptor‐mediated input. Population activity was greatly depressed by carbenoxolone, suggesting that propagation of depolarizing synaptic GABAergic potentials may be a critical determinant of interneuronal synchronous bursting in the hippocampus.