GABAergic cell types and modulation of GABAergic synaptic transmission by group III metabotropic glutamate receptors in the human neocortex

The neocortex is a layered, modular structure on the surface of the mammalian brain, which together with other brain structures supports behavioural adaptation to the changing environment. It is greatly expanded in humans and is thought to be necessary for unique human behaviours such as abstraction and language, functions which contribute to the high flexibility of human behaviour and underlie the adaptability of the species.
In the neocortex, the activity of spiny pyramidal neurons correlates with behaviourally relevant information processing and is governed by multiple types of GABAergic interneuron, which show great diversity and specialisation in the distribution of their dendrites and axon, molecular composition, electrical behaviour and synaptic connections. Although some GABAergic neuron types have been extensively studied in non-human animals, other cell types, such as the double bouquet cell (DBC) have been less frequently reported. Double bouquet cells are abundant in humans in layer II and upper layer III with a radial ‘horsetail’ axon descending through multiple layers in a narrow cortical column, yet details of their molecular content, electrical behaviour and synaptic targets in humans are not known.
The synaptic signalling of different types of GABAergic interneuron is regulated by presynaptic receptors, including group III metabotropic glutamate receptors (mGluR). In rodents, group III mGluRs show a cell type-specific expression in the neocortex and inhibit neurotransmitter release presynaptically, including that of GABA. However, their distribution and effects in human neocortex are not known. Although, drugs acting on presynaptic group III mGluRs have been proposed and investigated for treatment of various mental and neurodegenerative disorders in humans, none have been introduced into clinical practice so far. A direct investigation of group III mGluR function in human neocortex may help explain their effects, or the lack thereof, in health and disease and could facilitate development of more efficacious drugs.
The aims of my studies have been twofold: i, to explore the diversity of GABAergic interneuron types in the human neocortex by anatomical, molecular and electrophysiological methods, and ii, to test if GABAergic synaptic transmission is suppressed by group III mGluR activation in the human neocortex.
I recorded and labelled single and pairs of GABAergic interneuron, using the whole-cell patch clamp technique in vitro in acute neocortical slices prepared from samples obtained from tumour and epilepsy patients undergoing neurosurgery. I tested the effects of group III mGluR activation on spontaneous GABAergic synaptic transmission in two distinct GABAergic cell types, DBCs and parvalbumin-expressing multipolar cells (PMCs) by bath application of the orthosteric group III mGluR agonist, L-amino-phosphonobutyric-acid (L-AP4). I studied and compared the molecular expression and synaptic targets of DBCs and PMCs using immunohistochemistry and electron microscopy.
My main results are the following: 1. I identified 5 different GABAergic interneuron types based on the distribution of their dendrites and axon, including DBCs, PMCs, axo-axonic cells (AACs), neurogliaform cells (NGCs) and rosehip cells (RHCs). 2. The calcium-binding proteins calbindin (CB), calretinin (CR) and parvalbumin (PV) were differentially expressed among DBCs, and PMCs, such that the majority of DBCs were immunoreactive for CB and a small proportion were immunoreactive for CR, whereas the majority of PMCs were immunoreactive only for PV. 3. Voltage responses of DBCs were characterised by a large voltage sag potential and strong adaptation, whereas those of PMCs showed small or no voltage sag potential and weak or no adaptation. 4. Evoked firing patterns of DBCs exhibited bursting or non-bursting phenotypes. 5. Evoked firing patterns of DBCs and PMCs did not depend on medical history and did not vary with cortical area. 6. Half of the synaptic targets of the ‘horsetail’ axon of CB+ DBCs in layer III were dendritic spines, the remainder were dendritic shafts. In the neuropil of layer III, on average, 15% of GABAergic synapses innervated dendritic spines, the rest dendritic shafts; therefore, targeting of dendritic spines by DBCs is selective. 7. The majority (83%) of the synaptic targets of the CR+ DBC were dendritic shafts, the remainder dendritic spines. Two thirds of the synaptic targets of PMCs were dendritic shafts, the remainder dendritic spines and to a smaller extent somata. 8. Postsynaptic effects of DBCs were outward currents and showed short-term depression on their synaptic targets, which included pyramidal cells, DBCs, bipolar and bi-tufted smooth-dendritic neurons. 9. Activation of group III mGluRs paradoxically increased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) in DBCs, but decreased sIPSC frequency in PMCs; there was no effect on the amplitude of sIPSCs in either cell type.
My results reveal the differences in the molecular content, electrical behaviour and synaptic targets of DBCs and PMCs, and the cell type-specific effects of group III mGluR activation on GABAergic synaptic transmission onto these two distinct cell types in humans. I demonstrate a paradoxical enhancement of spontaneous GABAergic synaptic input to DBCs, which has not been observed in any species and is most likely due to a network effect. These results could provide experimental basis for the interpretation of drug action in the human cerebral cortex and for the development of novel treatments. Furthermore, I provide an integrated anatomical, molecular and electrophysiological characterisation of human DBCs, as an example of how to distinguish different cell types in the neocortex of humans and other species.

Keywords: presynaptic receptor
L-AP4
GABA-A receptor
inhibition
cell type
Advisor: Somogyi, P
Akerman, C