The downstream signaling of glutamate receptors that influence learning and memory can also be affected by cannabis exposure. Earlier, we discussed that once glutamate binds to the AMPA receptors, the postsynaptic membrane is depolarized, followed by the activation of NMDA channels. Only then do NMDA channels allow the entry of Ca²⁺ ions, which ultimately activate the second messenger system to aid in the formation of memory. Some other sources also supply Ca²⁺, like the endoplasmic reticulum. The endoplasmic reticulum releases calcium through the ryanodine receptor-sensitive calcium channel to increase the supply of intracellular calcium in the post-synaptic neuron. Chronic activation of cannabis receptors by CB₁ agonist WIN 55,212-2 decreased the supply of intracellular calcium, resulting in the alteration of the second messenger system’s activity [77]. However, when treated acutely, the cannabinoid agonist HU-210 increased intracellular calcium [78]. Following calcium entry, a kinase enzyme called protein Kinase C (PKC) is activated by calcium. During the long-term storage of memory and LTP, PKC-dependent phosphorylation of NMDARs causes the activation of NMDARs and helps to express more AMPARs in the synaptic surface [79]. PKC has several neuron-specific substrates required for learning and the formation of memories [80], among which neurogranin is one of the proteins located post-synaptically. Neurogranin modulates the binding and unbinding of the calcium-binding protein calmodulin (CaM), which controls calcium binding and leads to memory formation [81,82]. Acutely activated CB₁ receptors stimulate the phosphorylation of PKC and the phosphorylation of neurogranin, leading to no availability of free CaM for binding with Ca²⁺ to help in memory formation. This is one of the proposed mechanisms for how the acute activation of CB₁ interferes with short-term memory formation [83]. Likewise, protein kinase A (PKA) is inhibited by acute exposure to CB₁ agonists in cell culture [84]. Thus, chronic cannabis exposure can affect memory by decreasing intracellular calcium, which eventually affects the second messenger system crucial for memory formation. Conversely, acute exposure induces transient alterations that may initially modulate synaptic plasticity but ultimately interfere with memory consolidation processes. This highlights the intricate interplay between cannabinoid receptor activation, calcium signaling, and kinase activity in shaping the effects of cannabis on memory function.

Central to the downstream signaling of NMDARs are the Mitogen-activated protein (MAP) kinases [85]. The three most important MAP kinases signaling in the central and peripheral nervous systems are the extracellular signal-related kinases (ERKs), p38 MAP kinases, and C-Jun N-terminal kinases [85,86]. It was already established from prior research that endogenous cannabinoids activate the MAP kinase signal transduction pathway [87]. Transfection of Chinese hamster ovary (CHO) cells with CB₁ receptors, followed by treatment of these cells with three CB₁ agonists, CP 55,940, THC, and WIN 55,212-2, revealed that all activated the MAP kinases [88]. In addition to that, the agonists showed dose dependency while activating ERK1/2, and the relative ranking of the three different agonists was as follows: CP 55,940 > THC > WIN 55,212-2 [88]. THC also activated the MAPK/ERK in the dorsal striatum and the nucleus accumbens of treated mice [89]. In human astrocytoma cells, acute exposure to the CB₁ agonist HU-210 induced ERK activation, which was the downstream consequence of Gi dissociation and PI3K/PKB activation [90]. Several other research labs showed, using different cell lines, that PI3K/PKB is needed to activate ERK. Apart from that, it was also shown that inhibition of adenylyl cyclase, followed by the inhibition of PKA, is one of the predominant pathways for ERK activation due to CB₁ stimulation [84,91,92,93]. In the hippocampus, THC also activated ERK [94]. But, in the hippocampus, the ERK activation was not PI3K/PKB dependent. Instead, it was dependent on cAMP. Also, THC induced the expression of several genes (c-Fos protein, Zif268, and BDNF mRNAs) following the activation of ERK [94]. In the caudate putamen and cerebella of the rats, acute exposure to THC increased ERK even though there was no change in the hippocampus, nucleus accumbens, and prefrontal cortex with the same acute dose. However, when the same animal was exposed chronically, homeostatic adaptation led to no ERK change in the caudate putamen and cerebellum but significant activation in the hippocampus and cerebellum. Also, the role of Ras-Grf1 in this dose-dependent ERK activation and adaptation was found, as there was no effect on ERK when Ras-grf1 knockout mice were used [95]. In the cerebral frontal cortex, acute exposure to the CB₁ agonist WIN increased c-raf, ERK, MEK, and pERK, yet, when administered chronically for five consecutive days, there was no effect due to desensitization of the CB₁ receptors [96]. However, in cultured neuronal stem cells derived from mice, the CB₁ agonist arachidonoyl-2-chloroethyl amide (ACEA) promotes neuronal maturation and differentiation by inhibiting the ERK1/2 pathway [97]. Acute exposure to the CB₁ agonist cannabidiol at two different concentrations (5 μm and 10 μM) phosphorylated Erk ½, though the phosphorylation was higher at 5 μm than 10 μm [93]. These different effects in different models (in vitro and in vivo) suggest that the cannabinoid-induced effect of ERK acts differently in various models and treatments.

In summary, it appears that the relationship between the cannabinoid receptors and the MAP kinase signaling pathway is complicated. While both endogenous cannabinoids and various CB₁ agonists consistently activate MAP kinases, the specific mechanisms vary across different cell types, brain regions, and experimental conditions. These cannabinoids exhibit diverse effects on ERK activation, from promoting neuronal maturation and differentiation to modulating gene expression and adaptation, both in vitro and in vivo. All these diverse effects underscore the importance of considering the specific context and conditions when studying cannabinoid-mediated signaling, highlighting the need for further research to elucidate the precise mechanisms and therapeutic implications of cannabinoid-induced ERK activation.

Apart from ERK, another essential part of the MAPK signaling pathway is p38 MAPK. In the hippocampus, when LTP is induced, p38 MAPK is activated [98], while inhibition of p38 MAPK protects the neuron from over-excitation from glutamatergic neurotransmission [99]. In terms of the effect of CB₁ receptors on p38 MAPK, incubation of both rat and mouse hippocampi with exogenous as well as endogenous CB₁ receptor agonists has been shown to increase the phosphorylation of p38 MAPK [100]. A brief exposure (5 min) of CHO cells to 1 μM of THC activated p38 and JNK activity [101], whereas prolonged exposure (6 h) of AF5 cell lines to 3 μM of THC led to inhibition of p38 MAPK activation [102]. Cannabidiol was also found to inhibit p38 when examined in PC12 cells cultured for 36 h with cannabidiol at 10⁻⁶ to 10⁻⁴ M concentrations [103]. Treatment of mice with THC for seven consecutive days caused phosphorylation of p38 MAPK [45]. Numerous factors, from variations in cell types to the utilization of animal models, cannabinoid exposure duration, and concentrations, coupled with the distinct pharmacological properties of THC and cannabidiol, contribute to the observed differences in these responses. Thus, in summary, p38 MAPK and phosphorylation of p38 play a crucial role in glutamatergic neurotransmission, particularly in synaptic plasticity mechanisms like LTP, as inhibition of p38 can protect against over-excitation caused by glutamatergic neurotransmission. While in most cases, CB₁ receptor agonists induce phosphorylation of p38 MAPK in rat and mice hippocampi, THC exhibits varied effects on p38 activation, stimulating it in CHO cells but inhibiting it in the AF5 cell line, while cannabidiol suppresses p38 MAPK in PC12 cells. Overall, the involvement of p38 MAPK in glutamatergic neurotransmission and synaptic plasticity adds another layer of complexity to the interaction between cannabinoids and the MAP kinase signaling pathway. These findings highlight the intricate interplay between cannabinoids and p38 MAPK signaling, suggesting that the effects may be context dependent. Further research is needed to elucidate the precise mechanisms underlying these discrepancies and their potential implications for synaptic plasticity and neuronal function.

To summarize, the MAPK signaling pathway, particularly ERK and p38 MAPK, is integral to various neuronal processes, including synaptic plasticity and neurotransmitter regulation. The relationship between cannabinoids and MAP kinase signaling is complex and context-dependent, influenced by factors such as cell type, brain region, and treatment duration. Understanding these intricacies is essential for elucidating the therapeutic potential of cannabinoid-mediated signaling pathways and their implications for neurological disorders.

Another downstream signaling pathway of NMDA receptors is PI3K/Akt, and it plays neuroprotective roles against neurotoxicity [104,105]. Acute activation of the CB₁ receptor with THC has been found to activate the PI3K/Akt pathway, provide protection against neurotoxicity, and help in neuronal survival [106,107]. In this activation process, the CB₁ agonist THC phosphorylates Akt, which depends on a PI3K-dependent pathway and protects the neuron from external toxic stimuli like glutamate-mediated neurotoxicity. The downstream signaling target of PI3K/AKT is mTOR, which is a serine/threonine kinase, also called the mammalian target of rapamycin [108]. mTOR plays a crucial role in developing neurons and modifying synaptic strength [109]. Acute treatment of mice with THC phosphorylated mTOR and led to increased protein synthesis [110]. The increased synthesis of protein was confirmed using the protein synthesis inhibitor anisomycin. The increased protein synthesis was found to correlate with the amnesic-like effects of THC. When anisomycin was used, it improved the amnesic-like effects of THC. Also, the pharmacological blockade of mTOR improved the amnesic-like effects [111].

As all the pathways converge at the CREB pathway, the machinery that helps in the new protein synthesis for memory formation [112], the effect of the CB₁ agonist on the activity of CREB was also assessed. Two separate studies on male rats revealed that acute treatment with THC, at doses ranging from 2.5 to 10 mg/kg in one study [113] and at 15 mg/kg in another [95], increased CREB phosphorylation (pCREB) in the hippocampus, caudate putamen, and cerebellum [95,113]. However, the response to chronic THC exposure revealed a different pattern. Chronic treatment resulted in adaptations within the same brain regions, attenuating the changes seen during acute exposure. Interestingly, chronic THC exposure at a dose of 15 mg/kg activated CREB in the prefrontal cortex (PFC) only. Rubino et al. did not observe any deviations in cerebellar activity, contrasting with Casu et al.′s discovery of diminished pCREB expression in the same area post-prolonged THC exposure at 10 mg/kg [113]. Both investigations were limited to male rats. The contradiction in results could be attributed to several factors. Notably, Rubino’s protocol entailed a higher THC dosage (15 mg/kg) administered bi-daily for 6.5 days. In contrast, Casu’s regimen utilized a marginally lower dose (10 mg/kg), which was given once daily over four consecutive weeks. Moreover, chronic THC exposure at 10 mg/kg in mice reduced the expression of both total CREB and pCREB in the hippocampus [70], though they did not separate out the changes based on sex. The differences in the strain and species of the animals (rat vs. mice), as well as the dosage and duration of exposure, may have played a role in such discrepancies.

In summary, the impact of cannabinoid compounds on glutamatergic neurotransmission is intricate, as outlined in Figure 1 and Table 2. This review highlights that both acute and chronic exposure to cannabinoid compounds, particularly THC, can elicit a range of effects on glutamatergic signaling, which may, at times, elicit opposing effects. While acute exposure predominantly exhibits inhibitory effects, chronic exposure can lead to adaptive changes. Furthermore, these effects are contingent upon factors such as brain region, age, sex, and dosage regimen. To fully grasp the mechanisms by which cannabinoids influence glutamatergic neurotransmission in specific brain areas, further research is imperative (Table 3).