Anatomical studies of the dopamine system in the hypothalamus and the pituitary gland

The hypothalamus is a small, evolutionarily conserved brain region, necessary for our survival as individuals and as a species. It collects various sensory inputs, process them to maintain homeostasis and to overcome stressors, and generates outputs that affect the autonomic nervous system, the endocrine system and somatomotor behaviors. Energy metabolism, fluid balance, thermoregulation, sleep, aggression and reproduction are examples of functions under direct and indirect hypothalamic control. The neurochemical basis for these regulations involve different neurotransmitters and neuromodulators. The catecholamine dopamine is highly associated with various hypothalamic functions and behaviors, and has early on been shown to be present in intrinsic hypothalamic populations as well as incoming axon terminals. It acts on two types of receptors, excitatory D1-type and inhibitory D2-type, of which both have been reported to be expressed in the hypothalamus. To increase our understanding of these circuitries, this thesis aims to investigate the dopamine system in the hypothalamus, and the structures closely related to its inputs and outputs, namely the circumventricular organs and the pituitary. Immunohistochemical methods were used to generate a comprehensive distribution map of dopamine’s two main receptors, D1 and D2, and the neurochemical identity of these dopamine-receptor expressing cells were characterized. While the D2 receptor was widely expressed, D1 expression was found to be sparse. The suprachiasmatic nucleus, however, showed the contrary expression pattern. The D2 receptor could be localized to parvocellular neurons as well as endocrine cells of the pituitary. Little evidence for dopamine receptor expression on the magnocellular neurons could, however, be observed. Evidence for D1 receptor expression was also found in the subcommissural organ and a sub-cluster of ependymal cells in the third ventricle. Tuberoinfundibular dopamine (TIDA) neurons, which release dopamine in the portal vessels and thereby inhibit lactotrophs and prolactin release, were investigated in greater detail with regards to modulatory input, and morphological features. Anatomical substrate for innervation by serotonin and hypocretin/orexin on TIDA cell body and dendrites was identified together with electrophysiological evidence for excitation and suppression by hypocretin/orexin and serotonin or selective serotonin reuptake inhibitors, respectively. Morphological studies of male mice and rat TIDA neurons were done on tissue section and marker filled neurons by means of immunohistochemistry. TIDA neurons were found to preferentially extend dendrites towards the third ventricle, possibly even into the ventricle. Axon terminals were found in the median eminence, but collateral branches oriented laterally could also be detected. An intermingling subcellular distribution of inhibitory and excitatory synapses, on somatic and dendritic level, was also identified. No significant differences could be observed in most morphological properties of mouse and rat TIDA neurons. However, rats exhibited a higher total number of TIDA neurons and a lower spine density than mice. Finally, the expression of three different calcium binding proteins, i.e. calbindin-D28k, calretinin and parvalbumin, were investigated within the arcuate nucleus. While both calbindin-D28k and calretinin could be detected in the arcuate nucleus, little evidence for parvalbumin expression was observed. None of the proteins were expressed in TIDA neurons or other investigated populations, except for proopiomelanocortin neurons that expressed both calbindin-D28k and calretinin. These neurons showed a rostrocaudal segregation of the two calcium binding proteins that resulted in two separate subpopulations. Overall, the studies presented in this thesis reveal a previously unappreciated abundance of dopaminergic involvement in the hypothalamic circuitries which will increase our understanding of mammalian homeostatic and endocrine control

Understanding the Central Nervous System Symptoms of Rotavirus : A Qualitative Review

This qualitative review on rotavirus infection and its complications in the central nervous system (CNS) aims to understand the gut-brain mechanisms that give rise to CNS driven symptoms such as vomiting, fever, feelings of sickness, convulsions, encephalitis, and encephalopathy. There is substantial evidence to indicate the involvement of the gut-brain axis in symptoms such as vomiting and diarrhea. The underlying mechanisms are, however, not rotavirus specific, they represent evolutionarily conserved survival mechanisms for protection against pathogen entry and invasion. The reviewed studies show that rotavirus can exert effects on the CNS trough nervous gut-brain communication, via the release of mediators, such as the rotavirus enterotoxin NSP4, which stimulates neighboring enterochromaffin cells in the intestine to release serotonin and activate both enteric neurons and vagal afferents to the brain. Another route to CNS effects is presented through systemic spread via lymphatic pathways, and there are indications that rotavirus RNA can, in some cases where the blood brain barrier is weakened, enter the brain and have direct CNS effects. CNS effects can also be induced indirectly as a consequence of systemic elevation of toxins, cytokines, and/or other messenger molecules. Nevertheless, there is still no definitive or consistent evidence for the underlying mechanisms of rotavirus-induced CNS complications and more in-depth studies are required in the future.Funding Agencies|Swedish Research CouncilSwedish Research CouncilEuropean Commission [2018-02862]</p

Prolactin Regulates Tuberoinfundibular Dopamine Neuron Discharge Pattern: Novel Feedback Control Mechanisms in the Lactotrophic Axis

Balance in the body's hormonal axes depends on feedback onto neuroendocrine hypothalamic neurons. This phenomenon involves transcriptional and biosynthetic effects, yet less is known about the potential rapid modulation of electrical properties. Here, we investigated this issue in the lactotrophic axis, in which the pituitary hormone prolactin is tonically inhibited by tuberoinfundibular dopamine (TIDA) neurons located in the hypothalamic arcuate nucleus. Whole-cell recordings were performed on slices of the rat hypothalamus. In the presence of prolactin, spontaneously oscillating TIDA cells depolarized, switched from phasic to tonic discharge, and exhibited broadened action potentials. The underlying prolactin-induced current is composed of separate low- and high-voltage components that include the activation of a transient receptor potential-like current and the inhibition of a Ca(2+)-dependent BK-type K(+) current, respectively, as revealed by ion substitution experiments and pharmacological manipulation. The two components of the prolactin-induced current appear to be mediated through distinct signaling pathways as the high-voltage component is abolished by the phosphoinositide 3-kinase blocker wortmannin, whereas the low-voltage component is not. This first description of the central electrophysiological actions of prolactin suggests a novel feedback mechanism. By simultaneously enhancing the discharge and spike duration of TIDA cells, increased serum prolactin can promote dopamine release to limit its own secretion with implications for the control of lactation, sexual libido, fertility, and body weight.</p

Rotavirus Downregulates Tyrosine Hydroxylase in the Noradrenergic Sympathetic Nervous System in Ileum, Early in Infection and Simultaneously with Increased Intestinal Transit and Altered Brain Activities

Previous studies have investigated the mechanisms of rotavirus diarrhea mainly by focusing on intrinsic intestinal signaling. Although these observations are compelling and have provided important mechanistic information on rotavirus diarrhea, no information is available on how the gut communicates with the central nervous system (CNS) during rotavirus infection or on how this communication initiates sickness symptoms. While rotavirus diarrhea has been considered to occur only due to intrinsic intestinal effects within the enteric nervous system, we provide evidence for central nervous system control underlying the clinical symptomology. Our data visualize infection by large-scale three-dimensional (3D) volumetric tissue imaging of a mouse model and demonstrate that rotavirus infection disrupts the homeostasis of the autonomous system by downregulating tyrosine hydroxylase in the noradrenergic sympathetic nervous system in ileum, concomitant with increased intestinal transit. Interestingly, the nervous response was found to occur before the onset of clinical symptoms. In adult infected animals, we found increased pS6 immunoreactivity in the area postrema of the brain stem and decreased phosphorylated STAT5-immunoreactive neurons in the bed nucleus of the stria terminalis, which has been associated with autonomic control, including stress response. Our observations contribute to knowledge of how rotavirus infection induces gut-nerve-brain interaction early in the disease. IMPORTANCE Previous studies have investigated the mechanisms of rotavirus diarrhea mainly by focusing on intrinsic intestinal signaling. Although these observations are compelling and have provided important mechanistic information on rotavirus diarrhea, no information is available on how the gut communicates with the central nervous system (CNS) during rotavirus infection or on how this communication initiates sickness symptoms. We show that rotavirus infection presymptomatically disrupts the autonomic balance by downregulating the noradrenergic sympathetic nervous system in ileum, concomitant with increased intestinal transit and altered CNS activity, particularly in regions associated with autonomic control and stress response. Altogether, these observations reveal that the rotavirus-infected gut communicates with the CNS before the onset of diarrhea, a surprising observation that brings a new understanding of how rotavirus gives rise to sickness symptoms.Funding Agencies|Swedish Research Council [2018-02862, 2020-06116]; Hjarnfonden [PS2021-0063]</p

Hypocretin/Orexin Peptides Excite Rat Neuroendocrine Dopamine Neurons through Orexin 2 Receptor-Mediated Activation of a Mixed Cation Current

Hypocretin/Orexin (H/O) neurons of the lateral hypothalamus are compelling modulator candidates for the chronobiology of neuroendocrine output and, as a consequence, hormone release from the anterior pituitary. Here we investigate the effects of H/O peptides upon tuberoinfundibular dopamine (TIDA) neurons – cells which control, via inhibition, the pituitary secretion of prolactin. In whole cell recordings performed in male rat hypothalamic slices, application of H/O-A, as well as H/O-B, excited oscillating TIDA neurons, inducing a reversible depolarising switch from phasic to tonic discharge. The H/O-induced inward current underpinning this effect was post-synaptic (as it endured in the presence of tetrodotoxin), appeared to be carried by a Na(+)-dependent transient receptor potential-like channel (as it was blocked by 2-APB and was diminished by removal of extracellular Na(+)), and was a consequence of OX2R receptor activation (as it was blocked by the OX2R receptor antagonist TCS OX2 29, but not the OX1R receptor antagonist SB 334867). Application of the hormone, melatonin, failed to alter TIDA membrane potential or oscillatory activity. This first description of the electrophysiological effects of H/Os upon the TIDA network identifies cellular mechanisms that may contribute to the circadian rhythmicity of prolactin secretion

Serotonin and Antidepressant SSRIs Inhibit Rat Neuroendocrine Dopamine Neurons:Parallel Actions in the Lactotrophic Axis

This work was supported by European Research Council ENDOSWITCH 261286, Swedish Research Council 2014-3906, Strategic Research Programme in Diabetes at Karolinska Institutet, and Novo Nordisk Fonden to C.B. D.J.L. was supported by Wenner-Gren Foundations postdoctoral fellowship. The authors thank Dr. April Johnston for advice on serotonin pharmacology and Dr. Pradeep Atluri for discussions on clinical manifestations of SSRI treatment.Peer reviewedPublisher PD

Viral Gastroenteritis: Sickness Symptoms and Behavioral Responses

Viral infections have a major impact on physiology and behavior. The clinical symptoms of human rotavirus and norovirus infection are primarily diarrhea, fever, and vomiting, but several other sickness symptoms, such as nausea, loss of appetite, and stress response are never or rarely discussed. These physiological and behavioral changes can be considered as having evolved to reduce the spread of the pathogen and increase the chances of survival of the individual as well as the collective. The mechanisms underlying several sickness symptoms have been shown to be orchestrated by the brain, specifically, the hypothalamus. In this perspective, we have described how the central nervous system contributes to the mechanisms underlying the sickness symptoms and behaviors of these infections. Based on published findings, we propose a mechanistic model depicting the role of the brain in fever, nausea, vomiting, cortisol-induced stress, and loss of appetite.Funding Agencies|Swedish Research Council [2018-02862, 2020-06116]; Hjarnfonden [PS2021-0063]; Region OEstergoetland [ALF ROE-969520]</p

Molecular interrogation of hypothalamic organization reveals distinct dopamine neuronal subtypes.

The hypothalamus contains the highest diversity of neurons in the brain. Many of these neurons can co-release neurotransmitters and neuropeptides in a use-dependent manner. Investigators have hitherto relied on candidate protein-based tools to correlate behavioral, endocrine and gender traits with hypothalamic neuron identity. Here we map neuronal identities in the hypothalamus by single-cell RNA sequencing. We distinguished 62 neuronal subtypes producing glutamatergic, dopaminergic or GABAergic markers for synaptic neurotransmission and harboring the ability to engage in task-dependent neurotransmitter switching. We identified dopamine neurons that uniquely coexpress the Onecut3 and Nmur2 genes, and placed these in the periventricular nucleus with many synaptic afferents arising from neuromedin S(+) neurons of the suprachiasmatic nucleus. These neuroendocrine dopamine cells may contribute to the dopaminergic inhibition of prolactin secretion diurnally, as their neuromedin S(+) inputs originate from neurons expressing Per2 and Per3 and their tyrosine hydroxylase phosphorylation is regulated in a circadian fashion. Overall, our catalog of neuronal subclasses provides new understanding of hypothalamic organization and function