During physiological ketosis, KB can supply almost 60% of the brain's energy requirement. Plasma KB pass the BBB through monocarboxylic acid transporters (MCT) in a gradient-dependent manner and are readily available for neurons and astrocytes (which are also ketogenic (1)). In fact, the brain is happy without glucose and using ketones (2). Besides being neuroprotective and metabolically more efficient, the main KB, bOHB (3-HB), could enhance memory and learning.Zou et al (3) tested this hypothesis in mice:"This study sought to investigate the effect of 3-HB and derivatives on neuroglial cell metabolic activity and gap junctional intercellular communication of hippocampal neurons, to evaluate the hippocampal expression of PUMA-G and proteins related to memory following treatment with 3-HB, and to determine whether 3-hydroxybutyrate methyl ester (3-HBME) improves learning and memory in the normal mouse."
Interneuronal communication can occur indirectly and directly. In the former, chemical synapses are involved, in which transmitters are released into the extracellular space and bind the postsynaptic cell membrane. In the latter, electrical synapses mediate communication. The most prevalent group of electrical synapses, neuronal gap junctions, connect directly the intracellular space of two cells by gap junction channels. Connexin proteins are the structural components of gap junction channels in the nervous system. Specifically, connexin 36 has been involved in learning and memory (4).3-HB is the endogenous ligand for PUMA-G (5). Its activation in adipose tissue is also produced by nicotinic acid, inhibiting lipolysis and controlling the rate of ketogenesis. This represents an homeostatic mechanism by which 3-HB controls its own production, preventing ketoacidosis. Researchers used the Morris water maze for measuring learning and memory, and analyzed hippocampal neuron exposure to 3-HB and derivatives in vitro. All of the metabolites stimulated metabolic activity in neuroglial cells. 3-HMBE increased gap junction intercellular communication, as well as connexin 36 expression by 30% (compared to 12% in mice treated with acetyl-L-carnitine) and pERK2 levels (phosphorylated ERK2, necessary for connexin assembly). Moreover, PUMA-G mRNA was found in the hippocampus, subthalamic nucleus, temporal cortex and frontal cortex, and 3-HBME enhanced the transcription of PUMA-G in the hippocampus.So this translates to improved memory and learning?"The escape latency of mice in all groups decreased with time. Overall treatment comparisons indicated that statistically significant differences existed among the groups. Notably, the 30 mM 3-HBME groups took less time (p < 0.05) than the control groups on days 1 and 3–5 to find the platform (Fig. 5B). All treatment groups were faster (p <0.05) than the control group on day 5."
Treatment groups include mice treated with either 3-HBME (20, 30 or 40mg/kg/d) or acetyl-L-carnitine. Control group was water. "Paths taken to the platform area on the fifth day of spatial training by mice in the 30 mg/kg/d 3-HBME group were more direct than those taken by mice in the control group, which took more circuitous paths."
Ketotic mice knew exactly what they wanted. This causes less total swimming distance (ie. increased efficiency):"Similar to the escape latency results, the total swimming distance of the 30 mM 3-HBME groups was shorter (p <0.05) than the control groups on days 1 and 3–5. (...) Moreover, the total swimming distance of mice in the 30 mg/kg/d 3-HBME group was shorter (p <0.05) than that of the control or other treatment groups at days 3–5."
A probe test* then was performed to evaluate memory. Three parameteres were measured: the number of crossings of the exact place where the platform had been located, the swimming distance in the quadrant of the former platform position, and the swimming path in the pool. Number of times that mice crossed the former position of the hidden platform within 60s: Swimming distance in the platform quadrant:Swimming path:Finally, thigmotaxis was lowest in the 30mg/kg/d 3-HMBE group. A retention test was performed two days after the probe test: These test showed that the 30mg/kg/d 3-HMBE group:a. Crossed more times the exact place where the platform had been located,b. Had the larger swimming distance in such quadrant,c. Found the platform faster than the other groups, d. Were the most calmed (less thigmotaxis), ande. Found the platform faster than the other groups during the retention test These results suggest that 3-HMBE, given at 30mg/kg/d, enhanced learning and memory**.Researchers used polyhydroxybutyrate (PHB) (a polyhydroxyalkanoate) to produce 3-HB and derivatives. Can we extrapolate these results to physiologically produced 3-HB? The finding that hippocampal neurons expressed PUMA-G receptors is encouraging. As mentioned, 3-HB is the endogenous ligand for PUMA-G, but there seems to be a desensitization of the receptor when exposed to large amounts of 3-HB compared to 3-HBME and 3-HBEE. As PUMA-G interaction with several ligands is not well understood, we cannot draw many conclusions. Some studies have evaluated the potential benefit of 3-HB in neurological disorders which compromise memory and learning, such as Alzheimer's disease (AD). For example, in some patients with AD and mild cognitive impairment, MCT oil produces an improvement on cognitive testing and paragraph recall (6), correlated with the increase in plasma ketones. This neuroprotective effect has been explained by the increase in metabolic efficiency associated with 3-HB (7) and inhibition of apoptosis (8), but it might also act through PUMA-G and connexin 36 dependent mechanisms. Further studies should help discovering the molecular pathways involved. In the meantime, maybe schools should start giving students coconut oil shots instead of skim milk. * The platform was removed from the pool and mice were challenged to a single search trial for 60s. ** Note that this group performed best than 20mg/kg/d and 40mg/kg/d.
Zou XH, Li HM, Wang S, Leski M, Yao YC, Yang XD, Huang QJ, & Chen GQ (2009). The effect of 3-hydroxybutyrate methyl ester on learning and memory in mice. Biomaterials, 30 (8), 1532-41 PMID: 19111894
Ketosis during pregnancy has been known for many years. Fetal growth depends on constant energy supply, so physiological mechanisms should have been developed during evolution to assure intra-uterine development under starvation or food scarcity. Most studies focusing on pregnancy and fetal development have been done, for obvious reasons, on animals. It is not possible to extrapolate every detail, but it gives us a great idea and explanation for the metabolic changes observed during pregnancy. Briefly, there are two metabolic periods clearly differentiated during gestation. The first one, corresponding to the first two thirds, is the anabolic phase characterized by hyperphagia and enhanced storage of body fat (we will call it Phase I). During the last third of gestation, the catabolic phase, fetal growth is very rapid, so the energy needs of the fetus are increased (1) (we will call it Phase II). Insulin metabolism, as an acquired evolutionary mechanism, plays a key role during this process. During Phase I, there is a 3.0 to 3.5 fold increase in first-phase and second-phase insulin release in response to glucose, without an alteration in peripheral IS (2). This assures that accumulation of protein, glucose and fat is appropriate for late pregnancy. As pregnancy progresses, this increase in glucose-stimulated insulin secretion is maintained, but IS is reduced in 50-70% (3, 4) during late pregnancy (Phase II). This mechanism serves to redistribute glucose and energy to the rapid growing fetus. In addition to peripheral IR (but not hepatic), gluconeogenesis (GnG) is increased 16 to 30% to supply the placenta and fetus demand. Contrary to the main GnG precursors in non-pregnant adults, glycerol is the main glucose precursor, which represents a mechanism by which in the abscence of food, the mother is capable of producing the necessary glucose from a substrate that is readily available during fasting and not depend on external substrates. This process is accentuated by fasting, commonly known as "accelerated starvation": compared to non-pregnant, women during gestation exhibit a pronounced hypoglycemia and rapid rise in KB. GnG increases parallels the rise in KB (4). Because of its increased utilization, glucose has drawn much attention away from the importance of KB in fetal development. bOHB is utilized in a dose-dependent manner by the rat conceptus (5) and serves to spare glucose and lactate for biosynthetic pathways (6). bOHB seems to be the main oxidative fuel to the human fetal brain, measured by the production of CO2 (7). A classic study done on rat embryos underscore the importance of both glucose and bOHB to a proper development (8). Researchers tested the effect of increasing doses of glucose, KB or both on organ teratogenesis. They first tested glucose alone. According to the authors:(...) we found that isosmotic supplementation of the culture medium with 12 mg/mL D-glucose during the 48-h incubations effected a generalized retardation of rat-embryo growth and lesions such as microencephaly, exencephaly, open neural tube, and pericardial edema (6). We documented specificity by demonstrating that the findings are not replicated with isosmotic equimolar additions of certain other hexoses, such as sorbitol, fructose, inositol, or galactose (6). Teratogenic potentialities of high glucose concentrations have also been demonstrated with cultured mouse embryos. Sadler elicited dysmorphogenic effects with increasing frequency by adding 5mg/mL or 8 mg/mL D-glucose to the suspending rat serum during mouse-embryo culture (33).
So high glucose concentrations are teratogenic for the embryo. They went further and examinated the effect of increasing doses. During the period of these studies in 1980-1981, isosmotic additions of 12 mg/ mL elicited a 49% incidence of minor and a 23% incidence of major lesions. By contrast isosmotic additions of 3 mg/mL D-glucose to the incubation medium did not evoke any discernible lesions during 48 h of culture, 6mg/mL resulted in only a 2.2% incidence of minor and no major lesions, and 9 mg/mL D glucose were required to elicit 5.1% major and 17.8% minor lesions in the cultured intact embryos from our outbred strain of Charles River Sprague-Dawley rats.
They concluded:(...) the dysmorphogenic potentialities of ambient glucose are clearly concentration dependent although the precise relationships may be quantitatively different in various species or in different strains from the same species.
So we know that hyperglicemia is teratogenic. But what about increasing doses of bOHB? Preliminary acute incubations with 14C-labelled 14C-hydroxybutyrate indicated that cultured embryo units can oxidize ketones on day 10.4 as well as 1 1.4 of development (36) so that ketones can subserve nutrient functions in some portions of the conceptus at both times. What about the effects of ketones on embryogenesis during these intervals? As summarized in Figure 3, isosmotic additions of 2 or 4 mM buffered D,L sodium (3-hydroxybutyrate during 48-h culture of rat conceptus from day 9.5 to 1 1 .5 of development did not elicit any discernible dysmorphogenesis.
So physiological concentrations of bOHB, as in a low carbohydrate diet, ARE NOT TERATOGENIC. Problems appear only when going above this threshold, as in DK. However, with 8 mM, 24.5% of the embryos developed minor lesions, and the inclusion of 16 mM D,L /3-hydroxybutyrate was associated with a 71% frequency of minor and 45% incidence of major lesions (36).
See the trend? With 8mM only a quarter developed minor lesions. But when levels went way up (not physiological) lesions are aggraviated.
On the left, added concentrations of glucose and on the right, added concentrations of bOHB. The trend is clear, there is no damage when KB are in the physiological range, but when levels increase to concentrations seen in DK, boom! As always, the problem arises with hyperketonemia, not ketosis. Its easier to develop hyperglycemia than hyperketonemia (except during starvation). Lastly, what happens if we mix the minimally teratogenic amount of glucose (6mg/dL) with the minimally teratogenic amount of bOHB (8mM)? Sinergy! 66% displayed minor lesions and 27.7% major lesions. Some of the effects could not be explained by normal growth retardation. KB are so important to normal growth that there is evidence that fetal ketogenesis occurs (9). To achieve an optimal development, the fetus must not be exposed to increased concentrations of KB nor glucose. Both sources of fuel are necessary but in the right amount. The body adapts to this situation increasing the production of glucose from glycerol, reducing the need for ingesting extra glucose. Increasing calories and carbohydrates during pregnancy predisposes the mother to hyperglycemia, GD and IR, neonatal macrosomy and teratogenesis. Reducing the GL of the diet has shown to offer benefits compared to a low-fat diet (10), even when carbohydrate intake is reduced to 40-45% of total calories (11). Controlled studies adressing the effects of less than 40% of carbohydrates are scarce (evil ketosis!). Nevertheless, going zero carb can be as dangerous as going high carb* (12). But there is no need to go up to 60%. In rats, the requirement for normal growth seems to be around 18-20% (13), comparable amount of carbohydrates eaten by most low carbers and/or paleo, while the human fetus consumes around 20-25g/glucose per day during late gestation (4). Maintaining a proper diet with plenty of saturated fat, low carbohydrate and adequate protein/EPA+DHA is essential for a healthy pregnancy. Quality over quantity. Any experiences to share?*Although this is physiolgically impossible. Only achievable eating zero carb protein drinks and oil.
