Neuronal networks which underlie Impaired Awareness of Hypoglycaemia

The counter-regulatory response (CRR) to hypoglycaemia requires behavioural, endocrine, and autonomic responses. The latter is important clinically, because sympathoadrenal activation produces symptoms, leading to hypoglycaemia awareness and corrective behaviours. A reduction in the efficacy of sympathoadrenal activation leads to impaired awareness of hypoglycaemia (IAH), a common condition which affects patients with insulin-treated diabetes and carries significant morbidity and mortality. These deficits are precipitated by repeated hypoglycaemia (RH), which can be experimentally modelled to gain mechanistic insights into IAH. The neurobiology which underlies this condition was investigated in this thesis using a mouse model of IAH. IAH possesses the hallmarks of habituation, which were recapitulated in our mouse model. This includes a replicable decrease in the response of the sympathoadrenal system to RH, response recovery in the absence of RH and response dishabituation with a heterotypic stimulus. We postulated that specific neuronal networks regulate habituation following RH, but in the context of diabetes and insulin therapy, this culminates in IAH. By using neuronal markers of acute activation and in vivo fibre photometry, corticotrophin-releasing hormone (CRH) neurons in the paraventricular nucleus of the hypothalamus (PVH) were identified as rapidly activated following acute stress, including hypoglycaemia and 2-DG-induced glucoprivation. In addition, the chemogenetic manipulation of CRH neurons in the medial parvocellular dorsal division of the PVH (CRHmpdPVH) confirmed that they are necessary for the full activation of the sympathoadrenal system following hypoglycaemia. Furthermore, neuronal tracing experiments indicated that CRHmpdPVH neurons connect with output elements of the sympathoadrenal system, through direct and indirect pathways involving CRH receptor-expressing neurons. Using in vivo fibre photometry, I also found that CRHmpdPVH neuronal activity habituated to the repeated homotypic stress of RH, but not to heterotypic stress, indicating regulation of stress responsivity by networks which encode stress familiarity. We postulated that these networks included neurons within the posterior paraventricular nucleus of the thalamus (pPVT), as RH was found to induce an increase in FosB, a marker of neuronal adaptation, in both the PVH and pPVT. Finally, using a genetically targeted ablation strategy, I demonstrated that glutamatergic pPVT neurons are necessary for the RH-induced habituation of the sympathoadrenal response. Furthermore, using monosynaptic silencing and neuronal tracing techniques, I demonstrated that the pPVT drives an inhibitory forebrain circuit, via the ventral bed nucleus of the stria terminalis (vBNST), which is functionally significant to sympathoadrenal response habituation. Combined, these findings localise RH-induced habituation to specific integrative neuronal networks, involving pPVT and CRHmpdPVH neurons. This circuit may be responsible for reduced sympathoadrenal activation following RH and potentially for the development of IAH.