Professor Ian Forsythe
Contact details
Tel: 0116 252 2922
Email: idf@le.ac.uk
Research
Molecular Neurophysiology Laboratory
We are interested in how brain cells (neurons) regulate their excitability and process information received from a sense organ, such as sound from the ear. Our research concerns mechanisms of hearing (and hearing loss or deafness) as employed in tasks such as extraction of a signal in noise and for localization of a sound source. We study how exposure to loud sound damages the ear and the brain, by inducing long term changes in gene expression and ion channel function. We have a particular interest on the Potassium Channel Family (there are over 80 genes specifying these channel subunits). We focus on the third family of voltage-gated potassium channels (known as ‘Kv3’), measuring their biophysical properties in identified neurons and their role(s) at the connections between neurons (at the synapse). We exploit the auditory brainstem to study synaptic transmission at the calyx of Held giant synapse (see below for details) and to explore pleotropic genetic defects in which mutation of a gene for an ion channel subunit (e.g. Kv3.3) may cause multiple physiological defects (for example causing movement and hearing disorders) as a means to understanding the fundamental roles of ion channels in these brain functions. These commonalities in the ways different parts of the brain work, allow our discoveries about hearing to apply more widely, informing us of general rules governing brain function and disease. For example, in epilepsy, stroke, ageing, ataxia, neurodegeneration and dementia.
Methods used: Whole cell patch recording, in vitro brain slice (brainstem, hippocampus, neocortex), voltage-clamp, calcium imaging, multi-electrode array, pharmocology, synaptic stimulation/transmission, immunohistochemistry, confocal microscopy, quantitative rtPCR, RNA sequencing, western blot, auditory stimulation and auditory brainstem response, CRISPR/cas9 gene editing.
Research Funding: BBSRC, MRC, EU-ITN Consortium, Rosetrees Trust, Autifony Therapeutics Ltd.
Laboratory Members:
Lab. Manager: Ms Michelle Hammett
Postdoctoral staff: Nasreen Choudhury
Postgraduate Students: Amy Richardson (BBSRC, CASE), Andrey Filipov (Leics Uni), Kseniia Bondarenko (EU LISTEN Consortium)
Alumni: Postdocs: Dr Sarah Lucas, Dr Beatrice Pigott, Dr Joshua Smalley, Dr Jonathan Roberts, Dr Conny Kopp-Scheinpflug, Sue Robinson, Dr Nadia Pilati, Dr Joern Steinert, Dr Tina Tong, Dr Tanya Chernova, Dr Rachael Hardman, Matt Barker, Dr Mike Postlethwaite, Dr Sarah Griffin, Dr Martine Hamann, Dr Brian Billups, Dr Euan Brown, Dr Adrian Wong, Dr Matt Cuttle, Dr Helen Brew, Dr Margaret Barnes-Davies.
Alumni: Postgrads: Dr Sherylanne Newton (2017), Dr Debora Linley (2017), Dr Julieta Campi (2016), Dr Marie Nugent (2015), Dr Jim Sinclair (2014), Dr Adam Tozer (2013), Dr Martin Haustein (2011), Dr Jamie Johnston (2008), Dr Paul Dodson (2003), Dr Steve Owens (2003), Dr Amanda Smith (1998), Dr Joanne Doughty (1996).
Intercalated BSc Medical Students: Georgina Yan, Michael Leung, Haresh Selvaskandan, Rebecca Allfree, Jessica Sterenborg, Craig Sheridan, Fatima Osmani.
Collaborators: Len Kaczmarek (Yale), Nadia Pilati and Charles Large (Autifony), Rodrigo Quian Quiroga (CSN), Conny Kopp-Scheinpflug (Munich, Germany), Richard Evans (MCB), Bruce Graham (Stirling), Matthias Hennig (Edinburgh), Claire Gibson, Vincenzo Marra and Jaime McCutcheon (NPB).
| An image of the human brain, compared to the size of a mouse brain. |
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Details of Research Interests:
1. Auditory processing. Each of the nuclei in the auditory brainstem contribute to processing information received from both ears (binaural). We use voltage-clamp to measure the synaptic and voltage-gated currents in identified neurons of each nucleus. By studying how synaptic currents integrate with the intrinsic currents to generate action potential firing, we can elucidate physiological mechanisms of auditory processing. For example, the ability to detect very short gaps in sound requires precise offset firing in the superior paraolivary nucleus (SPN). This offset firing requires three complementary processes: 1. A large chloride ion gradient is generated by the outwardly directed chloride pump or co-transporter KCC2. 2. A large glycinergic synaptic response (IPSP) then activates the ‘hyperpolarization-activated non-specific cation channels’, IH. 3. This suppresses activity during the sound, but causes rebound firing of action potentials at the end of a sound (Kopp-Scheinpflug et al., 2011) and so signals the ‘silence’ or end of a sound to higher levels of the brain. This enables gap-detection which is essential for vocal communication and language.
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The auditory brainstem receives information from both ears and computes the location of a sound – this is essential to escape from predators or to locate prey. Three of the key nuclei are labelled: LSO – lateral superior olive; SPN – superior paraolivary nucleus; MNTB – medial nucleus of the trapezoid body. |
Background - the Brainstem Auditory Pathway
Sound is detected by the inner hair cells in the cochlea, which excite the spiral ganglion cell processes so that bursts of action potentials pass into the brain along the 8th Nerve and project into the Cochlear Nucleus. Bushy cells of the anterior Ventral Cochlear Nucleus (aVCN) give a large projection which crosses the midline and forms the giant synapse or Calyx of Held in the contralateral Medial Nucleus of the Trapezoid Body (MNTB) which is mediated by the neurotransmitter glutamate. So what does the calyx of Held synapse with the MNTB do? It provides an inhibitory glycinergic projection to three key nuclei in the Superior Olivary Complex (SOC): The Medial Superior Olive (MSO); which computes interaural timing differences (ITD). The Lateral Superior Olive (LSO); which integrates an ipsilateral excitatory input with the inhibitory projection from the MNTB - to compute interaural level (or volume) difference (ILD). The MNTB also provides a powerful inhibition to the Superior Paraolivary Nucleus (SPN) which is concerned with detecting the end of a sound or gaps in sounds (see research projects). The common feature of auditory processing in this region is fast and accurate transmission of information as action potential trains. This enhances comparison of sound between both ears for the physiological functions of sound localisation in auditory space, for feature extraction in a noisy environment and for computation of temporal features, such as gap detection.
2. Hearing loss – Disease mechanisms in the auditory pathway. It is well established that loud sounds cause deafness by damaging hair cells in the cochlea; but extreme activation also triggers plasticity in the brains at the synapses and neurons of the auditory pathway and this can damage the brain itself. For example, we have recently shown that loud sounds change the expression of the glutamate receptor subunit genes, so as to favour synaptic channels with slower gating. We have also conducted two different types of experiment to examine the mechanisms of deafness. Sherylanne Newton is examining the changes in gene expression (RNA sequencing) following exposure to damaging levels of sound. We hope this will give us insights into changes associated with ageing and hearing loss. Second, Drs Emanuele Schiavon and Josh Smalley are studying how accumulation of bilirubin (such as happens in jaundice or with liver damage) causes deafness and demonstrated that this is associated with synaptic damage and induction of inflammatory signaling in the brain.
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Hair cells are aligned along the length of the cochlea to sense sound. |
3. The calyx of Held. This is the largest synapse in the brain and we developed the means to make direct recordings from it (Forsythe, 1994); this allowed the study of presynaptic calcium channels and transmitter release (exocytosis) at this glutamatergic synapse. In collaboration with Dr Vincenzo Marra we are examining vesicle recycling at the calyx of Held and it’s modulation by presynaptic mechanisms. Dr Sarah Lucas is using simultaneous presynaptic and postsynaptic recording to examine how differences in metabolic substrates (such as glucose and lactate) influence transmission at the calyx. This has implications for understanding the metabolic constraints on information transmission at all synapses in the brain. This is also part of a wider collaboration with computational neuroscientists Drs Bruce Graham, Matthias Hennig and Christophe Michel.
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The Calyx of Held is shown enclosing a single MNTB neuron. All three images are from one synapse (scale bar is 10 micrometers); the left and the right images are projections of the upper and lower halves of the giant synapse. The middle image is a single optical section through the ‘equator’: the ‘hole’ in the middle in the unstained MNTB neuron which is receiving input from one giant synapse (coloured red). |
Background - the Calyx of Held: a model excitatory synapse
This giant synapse was first discovered by the German anatomist Hans Held, who studied in Leipzig and first published his anatomical study of the central auditory pathway in 1893. One Hundred years later we were attempting to make electrophysiological recordings from this living synapse using an in vitro preparation.
4. Potassium channels and intrinsic excitability. We have a long interest in understanding the physiological role of different voltage-gated potassium channels and in explaining why there are more than 80 potassium channel subunit genes. Our recent work has focused on Kv2 and Kv3 families. Kv2 is the major delayed rectifier of the neocortex and is highly clustered and controlled by phosphorylation. Kv3 is responsible for generating very short action potentials; we have examining which subunit genes are expressed in the auditory brainstem and their role in auditory processing. Both Kv2 and Kv3 channels are modulated by nitric oxide (NO, which is generated by synaptic activity). NO suppresses Kv3 channels and facilitates Kv2, thereby switching the basis of action potential repolarization in the MNTB (Steinert et al., 2011). These modulatory mechanisms are thought to underlie the control of neuronal excitability – by a process called ‘intrinsic plasticity’. This is where synaptic inputs up- or down-regulate the neuronal potassium currents, so as to harmonize excitability to the overall synaptic activity. We are studying these signaling mechanisms in the brainstem and neocortex, where these processes are postulated to underlie pathological processes such as tinnitus and stroke.
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The auditory brainstem expresses different combinations of potassium channel genes in each nucleus. Here the image shows fluorescent antibodies labelling the potassium channel subunits Kv3.1 (in green) and Kv2.2 (in red). |
