Theta oscillations in the rat MS/DB complex in vitro: a locally generated, inhibition-based brain rhythm

The medial septum/diagonal band (MS/DB) complex, which forms part of the septohippocampal feedback loop, is thought to be critical for the generation and/or maintenance of the hippocampal theta rhythm (4-15 Hz) in vivo. Previously we demonstrated the presence of theta frequency rhythmic activity, within an in vitro slice preparation of the MS/DB, following application of the AMPA/kainate receptor agonist kainate (25–100 nM). Here we further investigate this rhythmic activity, in particular the effects of GABAA receptor mediated inhibition, on kainate induced theta activity in the MS/DB, using bath application of bicuculline. Longitudinal slices of the MS/DB (450 μm), from 21 day old rats, were maintained at 32°C in an interface recording chamber perfused with oxygenated ACSF. Following the bath application of kainate at 100 nM to the MS/DB, extracellular recordings of neuronal population activity, using ACSF-filled micropipettes, revealed rhythmic theta frequency activity. Bicuculline was then applied to the recording chamber and subsequent changes to the kainate induced activity were recorded at 30 and 60 minutes there after. Application of 10 µM bicuculline produced a significant reduction (P< 0.05, Student's T test) in the activity recorded at theta frequency (spectral integral in 4-15 Hz range) and in peak amplitude within 60 minutes of application (n=6). These results indicate a pivotal role for inhibitory transmission in pacing and maintaining the rhythmic activity that is observed in the MS/DB slice preparation following kainate induced theta frequency oscillations. SPONSOR: British Neuroscience Association

Kainate induced theta-frequency oscillatory network activity in the medial septum/diagonal band complex

The medial septum/diagonal band complex (MS/DB) forms part of the septo-hippocampal feedback loop and is thought to have a major functional role in the generation and/or maintenance of the hippocampal theta rhythm in vivo (4 * 15 Hz). Several different mechanistic scenarios may underlie the generation of a theta-frequency EEG pattern, amongst them (1) an external pacemaker-type input (2) theta activity being an emergent property of the septo-hippocampal feedback loop and (3) theta arising in the synaptic network of the MS/DB itself. This investigation tested the latter scenario by using an in vitro slice preparation of the (deafferented) MS/DB. Longitudinal slices (0.45 mm) from 21 day old rats were maintained at 32 deg C in an interface recording chamber perfused with oxygenated ACSF. Following the bath application of the AMPA/kainate receptor agonist kainate (25 *100 nM), extracellular recordings, using ACSF-filled micropipettes, showed rhythmic population activity with a mean peak frequency of ~6 Hz which was most prominent along the midline of the MS/DB. The higher concentrations of kainate were accompanied by corresponding increases in spectral power (amplitude). Subsequently, intracellular recordings were obtained with QX-314 containing electrodes to prevent spiking-activity, and thus allowing IPSPs to be recorded at depolarised membrane potentials. These recordings revealed the presence of rhythmic IPSPs (~6 Hz) in the class of fast-firing cells of oscillating MS/DB slices, presumably arising in the mutually connected interneuronal network of the MS/DB and pacing the oscillation. Moreover, these findings clearly demonstrate that the intrinsic circuitry of the isolated MS/DB complex is sufficient to generate rhythmic theta frequency activity

Kainate induced theta-frequency oscillatory network activity in the medial septum/diagonal band complex

The medial septum/diagonal band complex (MS/DB) forms part of the septo-hippocampal feedback loop and is thought to have a major functional role in the generation and/or maintenance of the hippocampal theta rhythm in vivo (4 * 15 Hz). Several different mechanistic scenarios may underlie the generation of a theta-frequency EEG pattern, amongst them (1) an external pacemaker-type input (2) theta activity being an emergent property of the septo-hippocampal feedback loop and (3) theta arising in the synaptic network of the MS/DB itself. This investigation tested the latter scenario by using an in vitro slice preparation of the (deafferented) MS/DB. Longitudinal slices (0.45 mm) from 21 day old rats were maintained at 32 deg C in an interface recording chamber perfused with oxygenated ACSF. Following the bath application of the AMPA/kainate receptor agonist kainate (25 *100 nM), extracellular recordings, using ACSF-filled micropipettes, showed rhythmic population activity with a mean peak frequency of ~6 Hz which was most prominent along the midline of the MS/DB. The higher concentrations of kainate were accompanied by corresponding increases in spectral power (amplitude). Subsequently, intracellular recordings were obtained with QX-314 containing electrodes to prevent spiking-activity, and thus allowing IPSPs to be recorded at depolarised membrane potentials. These recordings revealed the presence of rhythmic IPSPs (~6 Hz) in the class of fast-firing cells of oscillating MS/DB slices, presumably arising in the mutually connected interneuronal network of the MS/DB and pacing the oscillation. Moreover, these findings clearly demonstrate that the intrinsic circuitry of the isolated MS/DB complex is sufficient to generate rhythmic theta frequency activity

Involvement of electrical signalling in theta frequency oscillations generated in the medial septum/diagonal band of Broca in vitro

Theta frequency (4 – 15 Hz) extracellular field activity can be reliably and repeatedly evoked in the medial septum/diagonal band of Broca (MS/DB) in vitro by the application of kainate. We have previously demonstrated a critical role for inhibitory neurotransmission in the maintenance of this activity and presented data that indicates parvalbumin GABAergic neurons are responsible for pacing rhythmic theta activity in the MS/DB. Evidence of functional electrical coupling between GABAergic interneurons has been reported in the hippocampus and neocortex and it has recently become clear that gap junctions may have a significant function in the generation of neuronal population activity. Here we demonstrate that application of the gap-junction uncoupling agent carbenoxolone causes a significant reduction (P< 0.05, Paired T test) in the kainate induced activity recorded at theta frequency (spectral integral in 4-15 Hz range) in the MS/DB slice in vitro. MS/DB slices were prepared from male Wistar rats (21 days), which were terminally anaesthetised with pentobarbitone sodium (120 mg.kg-1, i.p.). The animals were transcardially perfused with ~ 25ml of modified ACSF and rapidly decapitated. Longitudinal slices (450 µm) were placed in an interface recording chamber and maintained at 32°C. Persistent theta oscillations were induced by bath application of 100 nM kainate. Carbenoxolone was bath applied to stabilized kainate induced activity at a concentration of 100 μM for 60 minutes. At 60 minutes there was a significant reduction in the theta frequency activity recorded (56.6% ± 2.3) and in peak amplitude (n = 6). These results indicate a pivotal role for electrical signalling in the theta frequency rhythmic activity induced in the MS/DB upon application of kainate

Involvement of electrical signalling in theta frequency oscillations generated in the medial septum/diagonal band of Broca in vitro

Theta frequency (4 – 15 Hz) extracellular field activity can be reliably and repeatedly evoked in the medial septum/diagonal band of Broca (MS/DB) in vitro by the application of kainate. We have previously demonstrated a critical role for inhibitory neurotransmission in the maintenance of this activity and presented data that indicates parvalbumin GABAergic neurons are responsible for pacing rhythmic theta activity in the MS/DB. Evidence of functional electrical coupling between GABAergic interneurons has been reported in the hippocampus and neocortex and it has recently become clear that gap junctions may have a significant function in the generation of neuronal population activity. Here we demonstrate that application of the gap-junction uncoupling agent carbenoxolone causes a significant reduction (P< 0.05, Paired T test) in the kainate induced activity recorded at theta frequency (spectral integral in 4-15 Hz range) in the MS/DB slice in vitro. MS/DB slices were prepared from male Wistar rats (21 days), which were terminally anaesthetised with pentobarbitone sodium (120 mg.kg-1, i.p.). The animals were transcardially perfused with ~ 25ml of modified ACSF and rapidly decapitated. Longitudinal slices (450 µm) were placed in an interface recording chamber and maintained at 32°C. Persistent theta oscillations were induced by bath application of 100 nM kainate. Carbenoxolone was bath applied to stabilized kainate induced activity at a concentration of 100 μM for 60 minutes. At 60 minutes there was a significant reduction in the theta frequency activity recorded (56.6% ± 2.3) and in peak amplitude (n = 6). These results indicate a pivotal role for electrical signalling in the theta frequency rhythmic activity induced in the MS/DB upon application of kainate

Critical role for Piccolo in synaptic vesicle retrieval

Loss of function of the presynaptic active zone protein Piccolo has recently been linked to a devastating disease causing brain atrophy. Here, we address how Piccolo inactivation adversely affects synaptic function and thus may contributes to neuronal loss. Our analysis shows that Piccolo is critical for the activity dependent recycling and maintenance of synaptic vesicles (SVs). Specifically, we find that boutons lacking Piccolo have deficits in the Rab5/EEA1 dependent formation of early endosomes and thus the recycling of SVs. Mechanistically, impaired Rab5 function was caused by the reduced synaptic recruitment of Pra1, known to interact selectively with the zinc fingers of Piccolo. Importantly, over-expression of GTPase deficient Rab5 or the Znf1 domain of Piccolo restores the size and recycling of SV pools. These data provide a molecular link between the active zone and endosome sorting at synapses providing hints to how Piccolo contributes to both developmental and psychiatric disorders

ANCHOR: web server for predicting protein binding regions in disordered proteins

Summary: ANCHOR is a web-based implementation of an original method that takes a single amino acid sequence as an input and predicts protein binding regions that are disordered in isolation but can undergo disorder-to-order transition upon binding. The server incorporates the result of a general disorder prediction method, IUPred and can carry out simple motif searches as well

Treadmilling FtsZ polymers drive the directional movement of sPG-synthesis enzymes via a Brownian ratchet mechanism.

The FtsZ protein is a central component of the bacterial cell division machinery. It polymerizes at mid-cell and recruits more than 30 proteins to assemble into a macromolecular complex to direct cell wall constriction. FtsZ polymers exhibit treadmilling dynamics, driving the processive movement of enzymes that synthesize septal peptidoglycan (sPG). Here, we combine theoretical modelling with single-molecule imaging of live bacterial cells to show that FtsZ's treadmilling drives the directional movement of sPG enzymes via a Brownian ratchet mechanism. The processivity of the directional movement depends on the binding potential between FtsZ and the sPG enzyme, and on a balance between the enzyme's diffusion and FtsZ's treadmilling speed. We propose that this interplay may provide a mechanism to control the spatiotemporal distribution of active sPG enzymes, explaining the distinct roles of FtsZ treadmilling in modulating cell wall constriction rate observed in different bacteria