Cultivating meat from stem cells rather than by raising animals is a promising solution to concerns about the negative externalities of meat production. For cultivated meat to fully mimic conventional meat's organoleptic and nutritional properties, innovations in scaffolding technology are required. Many scaffolding technologies are already developed for use in biomedical tissue engineering. However, cultivated meat production comes with a unique set of constraints related to the scale and cost of production as well as the necessary attributes of the final product, such as texture and food safety. This review discusses the properties of vertebrate skeletal muscle that will need to be replicated in a successful product and the current state of scaffolding innovation within the cultivated meat industry, highlighting promising scaffold materials and techniques that can be applied to cultivated meat development. Recommendations are provided for future research into scaffolds capable of supporting the growth of high-quality meat while minimizing production costs. Although the development of appropriate scaffolds for cultivated meat is challenging, it is also tractable and provides novel opportunities to customize meat properties.

Patch-seq, combining patch-clamp electrophysiology with single-cell RNA-sequencing (scRNAseq), enables unprecedented access to a neuron's transcriptomic, electrophysiological, and morphological features. Here, we present a re-analysis of five patch-seq datasets, representing cells from ex vivo mouse brain slices and in vitro human stem-cell derived neurons. Our objective was to develop simple criteria to assess the quality of patch-seq derived single-cell transcriptomes. We evaluated patch-seq transcriptomes for the expression of marker genes of multiple cell types, benchmarking these against analogous profiles from cellular-dissociation based scRNAseq. We found an increased likelihood of off-target cell-type mRNA contamination in patch-seq cells from acute brain slices, likely due to the passage of the patch-pipette through the processes of adjacent cells. We also observed that patch-seq samples varied considerably in the amount of mRNA that could be extracted from each cell, strongly biasing the numbers of detectable genes. We developed a marker gene-based approach for scoring single-cell transcriptome quality post-hoc. Incorporating our quality metrics into downstream analyses improved the correspondence between gene expression and electrophysiological features. Our analysis suggests that technical confounds likely limit the interpretability of patch-seq based single-cell transcriptomes. However, we provide concrete recommendations for quality control steps that can be performed prior to costly RNA-sequencing to optimize the yield of high-quality samples.

In order to further our understanding of how gene expression contributes to key functional properties of neurons, we combined publicly accessible gene expression, electrophysiology, and morphology measurements to identify cross-cell type correlations between these data modalities. Building on our previous work using a similar approach, we distinguished between correlations which were “class-driven,” meaning those that could be explained by differences between excitatory and inhibitory cell classes, and those that reflected graded phenotypic differences within classes. Taking cell class identity into account increased the degree to which our results replicated in an independent dataset as well as their correspondence with known modes of ion channel function based on the literature. We also found a smaller set of genes whose relationships to electrophysiological or morphological properties appear to be specific to either excitatory or inhibitory cell types. Next, using data from PatchSeq experiments, allowing simultaneous single-cell characterization of gene expression and electrophysiology, we found that some of the gene-property correlations observed across cell types were further predictive of within-cell type heterogeneity. In summary, we have identified a number of relationships between gene expression, electrophysiology, and morphology that provide testable hypotheses for future studies.

Presynaptic neurexins (Nrxs) and type IIa receptortype protein tyrosine phosphatases (RPTPs) organize synapses through a network of postsynaptic ligands. We show that leucine-rich-repeat transmembrane neuronal proteins (LRRTMs) differentially engage the protein domains of Nrx but require its heparan sulfate (HS) modification to induce presynaptic differentiation. Binding to the HS of Nrx is sufficient for LRRTM3 and LRRTM4 to induce synaptogenesis. We identify mammalian Nrx1g as a potent synapse organizer and reveal LRRTM4 as its postsynaptic ligand. Mice expressing a mutant form of LRRTM4 that cannot bind to HS show structural and functional deficits at dentate gyrus excitatory synapses. Through the HS of Nrx, LRRTMs also recruit PTPs to induce presynaptic differentiation but function to varying degrees in its absence. PTPs forms a robust complex with Nrx, revealing an unexpected interaction between the two presynaptic hubs. These findings underscore the complex interplay of synapse organizers in specifying the molecular logic of a neural circuit.

Formation of synapses between neurons depends in part on binding between axonal and dendritic cell surface synaptic organizing proteins, which recruit components of the developing presynaptic and postsynaptic specializations. One of these presynaptic organizing molecules is protein tyrosine phosphatase σ (PTPσ). Although the protein domains involved in adhesion between PTPσ and its postsynaptic binding partners are known, the mechanisms by which it signals into the presynaptic neuron to recruit synaptic vesicles and other necessary components for regulated transmitter release are not well understood. One attractive candidate to mediate this function is liprin-α, a scaffolding protein with well-established roles at the synapse. We systematically mutated residues of the PTPσ intracellular region (ICR) and used the yeast dihydrofolate reductase (DHFR) protein complementation assay to screen for disrupted interactions between these mutant forms of PTPσ and its various binding partners. Using a molecular replacement strategy, we show that disrupting the interaction between PTPσ and liprin-α, but not between PTPσ and itself or another binding partner, caskin, abolishes presynaptic differentiation. Furthermore, phosphatase activity of PTPσ and binding to extracellular heparan sulfate (HS) proteoglycans are dispensable for presynaptic induction. Previous reports have suggested that binding between PTPσ and liprin-α is mediated by the PTPσ membrane-distal phosphatase-like domain. However, we provide evidence here that both of the PTPσ phosphatase-like domains mediate binding to liprin-α and are required for PTPσ-mediated presynaptic differentiation. These findings further our understanding of the mechanistic basis by which PTPσ acts as a presynaptic organizer.

In the original publication of this paper, the sentence ''The Lrrtm4RK-KI mouse model was designed and generated by the University of Manitoba Transgenic Facility

Cultivated meat, also known as cultured or cell-based meat, is meat produced directly from cultured animal cells rather than from a whole animal. Cultivated meat and seafood have been proposed as a means of mitigating the substantial harms associated with current production methods, including damage to the environment, antibiotic resistance, food security challenges, poor animal welfare, and—in the case of seafood—overfishing and ecological damage associated with fishing and aquaculture. Because biomedical tissue engineering research, from which cultivated meat draws a great deal of inspiration, has thus far been conducted almost exclusively in mammals, cultivated seafood suffers from a lack of established protocols for producing complex tissues in vitro. At the same time, fish such as the zebrafish Danio rerio have been widely used as model organisms in developmental biology. Therefore, many of the mechanisms and signaling pathways involved in the formation of muscle, fat, and other relevant tissue are relatively well understood for this species. The same processes are understood to a lesser degree in aquatic invertebrates. This review discusses the differentiation and maturation of meat-relevant cell types in aquatic species and makes recommendations for future research aimed at recapitulating these processes to produce cultivated fish and shellfish.

Antibiotic use in livestock and aquaculture production is driving resistance to medically important antibiotics. Producing meat through alternative methods, such as cultivated meat, offers an opportunity to decouple antibiotics from meat.