You will be presented with the abstract, introduction, and discussion section from a biomedical research article. Your task is to create a summary that a layperson can understand, capturing the essence of the research article. Keep essential scientific terms, but ensure that the language remains clear and the concepts are explained in an uncomplicated manner.

Title: Neurons are the Primary Target Cell for the Brain-Tropic Intracellular Parasite Toxoplasma gondii}
Article:
Abstract:
Toxoplasma gondii, a common brain-tropic parasite, is capable of infecting most nucleated cells, including astrocytes and neurons, in vitro. Yet, in vivo, Toxoplasma is primarily found in neurons. In vitro data showing that interferon-γ-stimulated astrocytes, but not neurons, clear intracellular parasites suggest that neurons alone are persistently infected in vivo because they lack the ability to clear intracellular parasites. Here we test this theory by using a novel Toxoplasma-mouse model capable of marking and tracking host cells that directly interact with parasites, even if the interaction is transient. Remarkably, we find that Toxoplasma shows a strong predilection for interacting with neurons throughout CNS infection. This predilection remains in the setting of IFN-γ depletion; infection with parasites resistant to the major mechanism by which murine astrocytes clear parasites; or when directly injecting parasites into the brain. These findings, in combination with prior work, strongly suggest that neurons are not incidentally infected, but rather they are Toxoplasma’s primary in vivo target.
Introduction:
Host cell-microbe interactions govern the survival and propagation of intracellular microbes. The importance of these interactions is particularly pronounced in chronic infections where persistence in specific cell types eludes pharmacologic or immunologic cures, leaving the pathogen to reactivate at opportunistic times. Understanding host cell-pathogen dynamics–from how pathogens find permissive cells to how they manipulate those cells–will provide opportunities to develop therapies that eliminate currently incurable persistent pathogens. Here, we seek to address how the ubiquitous intracellular parasite Toxoplasma gondii establishes a persistent brain infection. Toxoplasma is estimated to chronically infect the central nervous system( CNS) of up to 1/3 of the world’s population. This chronic CNS infection, for which there are no curative therapies, underlies Toxoplasma’s ability to reactivate and cause devastating neurologic disease and death in the immunosuppressed[1–3]. Prior to the introduction of effective combination anti-retroviral therapy, Toxoplasma was the most common cause of focal neurologic disease in AIDS patients[4]. Even today, toxoplasmic encephalitis occurs in untreated or undiagnosed AIDS patients as well as in patients on newer immunomodulants[3, 5–7]. Our understanding of Toxoplasma-CNS host cell interactions comes primarily from in vitro studies and the mouse model of toxoplasmosis. Like humans, mice are natural intermediate hosts in which the CNS is the major organ of encystment[8, 9]. Studies in human and rodent primary CNS cell cultures have established that Toxoplasma is capable of infecting and encysting in both astrocytes and neurons[10–12], two major parenchymal CNS cell types implicated in chronic infection. Yet, in vivo, Toxoplasma almost exclusively persists in neurons[13–15]. One of two alternative mechanisms could most easily reconcile these divergent in vitro and in vivo host cell preferences. One possibility is that both astrocytes and neurons are infected in vivo but only infected astrocytes either kill or are killed by the parasite, leaving neurons as the primary host cell for persistent infection. Alternatively, Toxoplasma could primarily interact with neurons in vivo, meaning that Toxoplasma persists in neurons because these are the cells with which the parasite predominantly interfaces. In support of the “astrocyte-killing” possibility, prior in vitro work established that stimulation with interferon-gamma( IFN-γ), the major cytokine required for systemic or cerebral resistance to Toxoplasma[16, 17], causes astrocytes, but not neurons, to use IFN-γ-regulated GTPases to kill up to 90% of the intracellular parasites[18–21]. While these studies imply that Toxoplasma primarily encysts in neurons because neurons cannot clear intracellular parasites, no studies have directly addressed this question in vivo, in part because of a lack of techniques to identify host cells with transient parasite interactions( e. g. transiently infected astrocytes that killed the intracellular parasite). We overcame this barrier by generating Toxoplasma strains that inject Cre recombinase into host cells[22]. Using these Toxoplasma-Cre strains to infect mice that express GFP in a Cre-dependent manner[23] enables the identification and tracking of host cells that directly interact with parasites. Importantly, this system causes a parasite-triggered permanent genetic change in host cells but does not require active infection. Therefore, host cells are permanently marked even in situations in which the host cell is injected with parasite proteins but is not invaded or in which the parasite invades and is subsequently killed by the host cell[24, 25]. Our prior work using this system revealed that at three weeks post-infection, the majority of the marked parenchymal CNS cells( GFP+ cells) appeared to be uninfected neurons[24]. As this finding was contradictory to what would be predicted by the “astrocyte-killing” model of CNS-Toxoplasma interactions, the work here expands upon those initial findings to more clearly define which parenchymal CNS cells directly interact with parasites in vivo. Remarkably, during CNS infection with two genetically divergent Toxoplasma-Cre strains, we find that neurons consistently make up the majority of the GFP+ parenchymal CNS cells regardless of time point or infecting strain. This neuronal predilection remains even in highly manipulated circumstances such as IFN-γ depletion or when infecting with parasites resistant to a major mechanism by which murine astrocytes eliminate intracellular parasites in vitro. Finally, imaging whole infected neurons in situ reveals that cysts are predominantly found in the extensive network of neuronal processes, not the neuron cell bodies. Collectively, these data highly suggest that Toxoplasma’s persistence in neurons is driven by a strong in vivo predilection for interacting almost exclusively with neurons and that physical properties such as the extensive size and breadth of neuronal processes may play a role in determining this in vivo preference.
Discussion:
In this study, we used a novel in vivo system to address an essential question: why does Toxoplasma gondii, a parasite with no clear host cell preference in vitro, show host cell preferences during in vivo infection of the CNS? Our ability to answer this question arose from the development of the Toxoplasma-Cre system, which is capable of identifying transient host cell-parasite interactions and allows imaging and analysis of entire infected neurons[22, 24, 42]. The data derived from this system and presented here strongly suggest that neuron-parasite interactions predominate throughout CNS infection and persist even in highly manipulated circumstances such as IFN-γ depletion, infection with IRG-resistant parasites, and direct intracranial inoculation of parasites. These data in combination with the recognition that cysts are most commonly found in distal neuronal processes indicate that during in vivo infection, Toxoplasma displays a distinct propensity for neurons, which may in part be driven by physical properties of the CNS and its cells. There are several important implications of these findings. Excitingly, the predominance of the neuron-parasite interaction implies that neurons are not mere bystanders that are incidentally infected by parasites, but rather that neurons are critical participants in the establishment of the persistent CNS infection, a concept consistent with a recent report showing that transgenic mice that lack gp130 in neurons are unable to control CNS toxoplasmosis[43]. In addition, the high number of uninfected GFP+ neurons[24] raises the tantalizing possibility that most neurons clear parasites by unrecognized means such as the recently described CD40-CD154 autophagy pathway[44, 45]. An equally provocative explanation for the uninfected GFP+ neurons is that direct parasitic manipulation of these long-lived CNS cells without invasion plays an unrecognized role in promoting parasite survival and transmission. In either case, future work will focus on determining the origins of the GFP+ uninfected neurons and how these neurons differ from actively infected neurons or neurons with no direct parasite interactions. Interestingly, our data from the intracranial injection experiments and our cyst location profiling suggest that physical properties of the CNS may play a role in driving this Toxoplasma-neuron predilection. As in vitro cultures cannot approximate the 3D nature of the in vivo CNS, these physical properties provide an appealing explanation for the discrepancy between prior in vitro work and our in vivo work. Of course, caution must be exercised when extrapolating from either set of experiments. The intracranial injection route is unnatural and skips the preceding systemic infection which provides cytokines that would be predicted to change the CNS environment even prior to parasite entry. The cyst location studies were done at a single time point, and do not address whether parasites invade and encyst in the same location or rather invade in one part of the neuron and migrate to the processes. Future work will focus on identifying the physical environment faced by parasites during entry into the CNS and how parasite location in the neuron changes from the time of entry to encystment. Of equal importance is the limited evidence we found for direct interactions between parasites and astrocytes. As noted before, the current understanding of astrocyte-Toxoplasma interactions is derived from in vitro work, and suggests that cysts are not found in vivo in astrocytes[15, 46] because astrocytes kill the intracellular parasites that readily invade this cell type[18, 20, 21, 33]. This “astrocyte-killing” model would predict that using our Toxoplasma-Cre system we should find many uninfected GFP+ astrocytes, but instead we observed few. The findings presented here offer a different explanation: cysts are seldom found in astrocytes because parasites rarely directly interact with astrocytes in hosts with intact IFN-γ responses. In addition, the lack of GFP+ astrocytes could be considered to indirectly support the possibility of parasites entering into the CNS via immune cell carriers[26, 47]. If parasites directly enter the CNS from the vasculature( either paracellularly or through infection of endothelial cells and egress from the basolateral side of the endothelial cell), parasites should next encounter astrocytes and the astrocytic endfeet that surround the vasculature[30]. Thus, to interact predominantly with neurons, parasites would have to migrate beyond the vasculature-associated astrocytes. On the other hand, if immune cells carry parasites into the CNS, parasite egress might only occur after infected immune cells pass these astrocytic processes. Of course, it is difficult to prove the non-existence of an interaction. Our system would not detect direct astrocyte-parasite interactions that occurred without the injection of parasite proteins, though such a phenomenon has yet to be described or predicted. Additionally, as noted above, our system would not detect direct astrocyte-parasite interactions if infected astrocytes died prior to the expression of GFP. We think astrocyte death is unlikely to account for the lack of evidence for direct astrocyte-parasite interactions for several reasons. First, while several classes of IFN-γ-dependent GTPases are used by astrocytes to kill intracellular parasites[20, 21], only IRG-deployment has been linked to subsequent death of the host cell in vitro[34]. We directly addressed this concern by using IRG-resistant III-Cre-ROP18 parasites and found no increase in the percentage of GFP+ cells identified as astrocytes( Fig 3). Second, given that we and others have reported significant numbers of GFP+ uninfected cells in vivo in immune cells[24, 25, 48] as well as neurons, rapid astrocyte death after parasite interaction would require invoking a unique mechanism of death that presumptively does not occur in these other cell types. Third, a recent report found no evidence for significant astrocytic death during CNS toxoplasmosis[41], further decreasing the likelihood that astrocyte death accounts for the paucity of GFP+ astrocytes. Thus, given the current lack of evidence for either parasite invasion without injection of parasite proteins or astrocytic death during CNS toxoplasmosis, these data suggest that astrocytes primarily constrain Toxoplasma growth through indirect mechanisms such as barrier formation and/or secretion of cytokines and chemokines[30], rather than by direct killing of parasites. This possibility is consistent with prior work showing that mice lacking the glial fibrillary acid protein( GFAP) or the gp130 receptor only in astrocytes–two defects that would not be expected to impact astrocytic killing of intracellular parasites–develop uncontrolled CNS toxoplasmic encephalitis[49, 50]. Additionally, the IFN-γ depletion experiments suggest that in severely immunocompromised patients, such as AIDS patients[51], an increase in direct parasite-astrocyte interactions may play an unrecognized role in the neuropathology of toxoplasmic encephalitis. Together, these findings offer new opportunities to dissect out the changing role of astrocytes in health and disease. In conclusion, the findings presented here strongly suggest a new paradigm for CNS-Toxoplasma interactions. In this new paradigm, direct neuron-Toxoplasma interactions play an active and central role in the establishment of a successful, chronic CNS infection.
Toxoplasma gondii is a ubiquitous intracellular parasite that chronically infects the brain of up to 1/3 of the world’s population . While most infected persons have no symptoms , Toxoplasma’s tropism for the brain underlies the parasite’s ability to cause debilitating neurologic disease in the developing fetus and the immunocompromised , such as AIDS or organ transplant patients . Our understanding of which cells of the brain interact with and are infected by Toxoplasma primarily comes from work in tissue culture , which cannot re-capitulate the complexity of a living animal . Here , we overcome this barrier by studying brain-Toxoplasma interactions using a novel mouse model that allows us to permanently mark and track brain cells that have interacted with parasites . Contrary to what has been shown in cell culture , we find that throughout acute and chronic brain infection Toxoplasma almost exclusively interacts with neurons , and rarely interacts with astrocytes . This neuronal predilection persists even in highly manipulated circumstances such as the introduction of parasites directly into the brain . This work is the first to show that Toxoplasma primarily interacts with neurons in vivo , which suggests that to understand Toxoplasma’s ability to persistently infect the brain , we must understand the neuron-Toxoplasma interaction .